Table of SynGAP1 Isoform α2 (UniProt Q96PV0-1) Missense Variants.

c.dna	Variant	SGM Consensus	Domain	ClinVar			gnomAD			ESM1b		AlphaMissense			REVEL		FoldX			Rosetta		Foldetta		PremPS		PROVEAN		PolyPhen-2 HumDiv		PolyPhen-2 HumVar		FATHMM		SIFT				PAM		Physical		SASA		Normalized B-factor backbone		Normalized B-factor sidechain		SynGAP Structural Annotation									DOI
				Clinical Status	Review	Subm.	ID	Allele count	Allele freq.	LLR score	Prediction	Pathogenicity	Class	Optimized	Score	Prediction	Average ΔΔG	Prediction	StdDev	ΔΔG	Prediction	ΔΔG	Prediction	ΔΔG	Prediction	Score	Prediction	pph2_prob	Prediction	pph2_prob	Prediction	Nervous System Score	Prediction	Prediction	Status	Conservation	Sequences	PAM250	PAM120	Hydropathy Δ	MW Δ	Average	Δ	Δ	StdDev	Δ	StdDev	Secondary	Tertiary bonds	Inside out	GAP-Ras interface	At membrane	No effect	MD Alert	Verdict	Description
c.2369C>A	T790N		SH3-binding motif	Conflicting		3	6-33442921-C-A	69	4.28e-5	-5.243	Likely Benign	0.276	Likely Benign	Likely Benign	0.103	Likely Benign										-2.54	Deleterious	0.999	Probably Damaging	0.997	Probably Damaging	2.27	Pathogenic	0.02	Affected	3.64	6	0	0	-2.8	13.00
c.2900G>A	R967Q	Likely Benign		Benign/Likely benign		2	6-33443452-G-A	31	1.92e-5	-3.057	Likely Benign	0.080	Likely Benign	Likely Benign	0.104	Likely Benign										-0.01	Neutral	0.994	Probably Damaging	0.626	Possibly Damaging	4.21	Benign	0.36	Tolerated	4.32	2	1	1	1.0	-28.06
c.1904A>G	N635S (3D Viewer)		GAP	Conflicting		4	6-33440956-A-G	10	6.20e-6	-9.002	Likely Pathogenic	0.101	Likely Benign	Likely Benign	0.104	Likely Benign	0.80	Ambiguous	0.1	0.67	Ambiguous	0.74	Ambiguous	0.95	Ambiguous	-4.45	Deleterious	0.261	Benign	0.044	Benign	3.06	Benign	0.05	Affected	3.37	34	1	1	2.7	-27.03	196.0	30.9	0.1	0.0	-0.3	0.2				X				Uncertain	In the WT simulations, the carboxamide side chain of Asn635, located on the outer surface of an α helix (res. Glu617-Asn635), forms hydrogen bonds with Gln631 on the same α helix and with the hydroxyl side chain of Ser590 on an opposing α helix (res. Glu582-Met603).In the variant simulations, the side chain of Ser635 is shorter than asparagine and thus prefers to hydrogen bond with the carbonyl group of Gln631 on the same helix and, to a lesser extent, with Ser590 compared to Asn635 in the WT. Ser635 forms hydrogen bonds with the backbone atoms of the same helix, which may destabilize the helix, although this is not clearly evident in the simulations. The weakening of the hydrogen bond between Ser635 and Ser590 in the variant may also weaken the tertiary structure assembly between the helices.Additionally, Asn635 is at the GTPase interface. However, the implication of the residue swap on the complex formation with the GTPase cannot be investigated using solvent-only simulations.
c.3209G>A	R1070K	Likely Benign		Conflicting		2				-5.093	Likely Benign	0.326	Likely Benign	Likely Benign	0.104	Likely Benign										-1.42	Neutral	0.049	Benign	0.048	Benign	3.86	Benign	0.09	Tolerated	3.77	5	3	2	0.6	-28.01
c.3314G>A	R1105Q	Likely Benign		Uncertain		2	6-33443866-G-A	3	1.96e-6	-3.666	Likely Benign	0.216	Likely Benign	Likely Benign	0.104	Likely Benign										-1.21	Neutral	0.958	Probably Damaging	0.194	Benign	2.50	Benign	0.16	Tolerated	3.77	5	1	1	1.0	-28.06
c.3846G>C	E1282D	Likely Benign		Uncertain		1	6-33447894-G-C	1	6.44e-7	-3.879	Likely Benign	0.074	Likely Benign	Likely Benign	0.104	Likely Benign										-1.26	Neutral	0.112	Benign	0.036	Benign	2.70	Benign	0.39	Tolerated	3.77	5	3	2	0.0	-14.03
c.3260C>T	S1087F			Uncertain		1				-3.843	Likely Benign	0.497	Ambiguous	Likely Benign	0.105	Likely Benign										-2.75	Deleterious	0.990	Probably Damaging	0.796	Possibly Damaging	2.56	Benign	0.03	Affected	3.77	5	-2	-3	3.6	60.10
c.44C>T	A15V	Likely Benign		Uncertain		1	6-33420308-C-T	1	6.49e-7	-3.560	Likely Benign	0.161	Likely Benign	Likely Benign	0.105	Likely Benign										0.20	Neutral	0.602	Possibly Damaging	0.015	Benign	4.19	Benign	0.00	Affected	4.32	1	0	0	2.4	28.05
c.3424T>C	S1142P	Likely Benign		Likely Benign		1	6-33444459-T-C	1	6.20e-7	-2.713	Likely Benign	0.222	Likely Benign	Likely Benign	0.107	Likely Benign										-2.19	Neutral	0.918	Possibly Damaging	0.761	Possibly Damaging	2.64	Benign	0.00	Affected	4.32	4	-1	1	-0.8	10.04
c.2573G>A	S858N	Likely Benign		Uncertain		1	6-33443125-G-A	2	1.24e-6	-4.311	Likely Benign	0.121	Likely Benign	Likely Benign	0.107	Likely Benign										-0.67	Neutral	0.448	Benign	0.846	Possibly Damaging	4.13	Benign	0.02	Affected	3.77	5	1	1	-2.7	27.03
c.2768T>A	I923N	Likely Benign		Uncertain		1				-0.733	Likely Benign	0.712	Likely Pathogenic	Likely Benign	0.108	Likely Benign										-1.16	Neutral	0.991	Probably Damaging	0.793	Possibly Damaging	2.70	Benign	0.13	Tolerated	3.77	5	-2	-3	-8.0	0.94
c.3172G>A	G1058S	Likely Benign		Conflicting		3	6-33443724-G-A	114	7.08e-5	-5.178	Likely Benign	0.081	Likely Benign	Likely Benign	0.108	Likely Benign										0.26	Neutral	0.001	Benign	0.001	Benign	5.38	Benign	0.04	Affected	3.77	5	1	0	-0.4	30.03
c.269T>A	V90E	Likely Benign		Uncertain		1				-4.079	Likely Benign	0.703	Likely Pathogenic	Likely Benign	0.108	Likely Benign										-0.38	Neutral	0.001	Benign	0.000	Benign	4.00	Benign	0.00	Affected	4.32	1	-2	-2	-7.7	29.98
c.416G>A	S139N	Likely Benign		Uncertain		1	6-33432713-G-A	3	2.22e-6	-4.584	Likely Benign	0.688	Likely Pathogenic	Likely Benign	0.109	Likely Benign										-0.75	Neutral	0.149	Benign	0.047	Benign	4.14	Benign	0.24	Tolerated	3.61	5	1	1	-2.7	27.03
c.2924C>G	T975S	Likely Benign		Uncertain		1				-2.743	Likely Benign	0.068	Likely Benign	Likely Benign	0.109	Likely Benign										-0.57	Neutral	0.059	Benign	0.061	Benign	4.16	Benign	0.20	Tolerated			1	1	-0.1	-14.03
c.3026A>C	E1009A	Likely Pathogenic		Uncertain		1				-3.118	Likely Benign	0.679	Likely Pathogenic	Likely Benign	0.109	Likely Benign										-3.06	Deleterious	0.980	Probably Damaging	0.630	Possibly Damaging	2.39	Pathogenic	0.01	Affected	3.77	5	0	-1	5.3	-58.04
c.3056G>T	R1019L	Likely Pathogenic		Uncertain		1	6-33443608-G-T	2	1.24e-6	-5.194	Likely Benign	0.752	Likely Pathogenic	Likely Benign	0.110	Likely Benign										-3.57	Deleterious	0.800	Possibly Damaging	0.573	Possibly Damaging	2.40	Pathogenic	0.01	Affected	3.77	5	-2	-3	8.3	-43.03
c.37A>G	I13V	Likely Benign		Uncertain		1				-2.497	Likely Benign	0.105	Likely Benign	Likely Benign	0.110	Likely Benign										0.01	Neutral	0.000	Benign	0.000	Benign	4.25	Benign	0.00	Affected			4	3	-0.3	-14.03
c.2623G>A	A875T	Likely Benign		Uncertain		1	6-33443175-G-A	1	6.20e-7	-3.793	Likely Benign	0.179	Likely Benign	Likely Benign	0.110	Likely Benign										-1.56	Neutral	0.972	Probably Damaging	0.864	Possibly Damaging	2.72	Benign	0.26	Tolerated	3.77	5	0	1	-2.5	30.03
c.505G>A	D169N			Uncertain		1				-10.713	Likely Pathogenic	0.761	Likely Pathogenic	Likely Benign	0.110	Likely Benign										-2.04	Neutral	0.079	Benign	0.052	Benign	4.07	Benign	0.01	Affected	3.74	4	2	1	0.0	-0.98
c.272A>G	E91G	Likely Benign		Likely Benign		1				-3.226	Likely Benign	0.783	Likely Pathogenic	Likely Benign	0.110	Likely Benign										-2.18	Neutral	0.947	Possibly Damaging	0.727	Possibly Damaging	3.86	Benign	0.00	Affected	4.32	1	0	-2	3.1	-72.06
c.2608C>G	L870V	Likely Benign		Uncertain		1				-4.123	Likely Benign	0.300	Likely Benign	Likely Benign	0.111	Likely Benign										-1.19	Neutral	0.997	Probably Damaging	0.992	Probably Damaging	2.64	Benign	0.12	Tolerated	3.88	3	2	1	0.4	-14.03
c.2047A>G	I683V (3D Viewer)	Likely Benign	GAP	Uncertain		1	6-33441306-A-G	2	1.24e-6	-7.588	In-Between	0.138	Likely Benign	Likely Benign	0.112	Likely Benign	0.90	Ambiguous	0.0	0.60	Ambiguous	0.75	Ambiguous	0.76	Ambiguous	-0.78	Neutral	0.538	Possibly Damaging	0.080	Benign	3.35	Benign	0.14	Tolerated	3.42	17	4	3	-0.3	-14.03	215.6	29.1	0.0	0.0	-0.7	0.1						X		Potentially Benign	The sec-butyl side chain of Ile683, located in an entangled α-α loop connecting the two α-helices (res. Ser641-Glu666 and res. Leu685-Val699), is sterically packed against His453 and Glu688. In the variant simulations, the iso-propyl side chain of Val683 has similar size and physicochemical properties as Ile630 in the WT, and thus, it is able to maintain similar interactions in the inter-helix space. Consequently, no negative structural effects are observed during the simulations due to the residue swap.
c.2724G>C	Q908H	Likely Benign		Conflicting		4	6-33443276-G-C	1	6.20e-7	-4.658	Likely Benign	0.311	Likely Benign	Likely Benign	0.112	Likely Benign										-0.74	Neutral	0.996	Probably Damaging	0.995	Probably Damaging	2.58	Benign	0.05	Affected	3.77	5	3	0	0.3	9.01
c.3002T>C	L1001P	Likely Benign		Uncertain		1				-3.071	Likely Benign	0.209	Likely Benign	Likely Benign	0.113	Likely Benign										-1.02	Neutral	0.966	Probably Damaging	0.690	Possibly Damaging	2.65	Benign	0.00	Affected	4.32	4	-3	-3	-5.4	-16.04
c.2459A>G	Y820C	Likely Pathogenic		Uncertain		1				-8.797	Likely Pathogenic	0.744	Likely Pathogenic	Likely Benign	0.113	Likely Benign										-3.16	Deleterious	1.000	Probably Damaging	0.983	Probably Damaging	2.68	Benign	0.06	Tolerated	3.77	5	0	-2	3.8	-60.04
c.2719A>T	S907C	Likely Benign		Likely Benign		1				-6.685	Likely Benign	0.298	Likely Benign	Likely Benign	0.113	Likely Benign										-2.34	Neutral	0.999	Probably Damaging	0.988	Probably Damaging	2.60	Benign	0.02	Affected	3.77	5	0	-1	3.3	16.06
c.335G>C	G112A	Likely Benign		Uncertain		1	6-33432200-G-C	15	9.30e-6	-2.456	Likely Benign	0.119	Likely Benign	Likely Benign	0.114	Likely Benign										-2.34	Neutral	0.231	Benign	0.054	Benign	4.07	Benign	0.00	Affected	3.61	5	1	0	2.2	14.03
c.3250C>G	P1084A	Likely Benign		Uncertain		1				-3.928	Likely Benign	0.066	Likely Benign	Likely Benign	0.114	Likely Benign										-2.54	Deleterious	0.649	Possibly Damaging	0.157	Benign	4.05	Benign	0.35	Tolerated	3.77	5	-1	1	3.4	-26.04
c.3943T>C	W1315R	Likely Benign		Uncertain		1				0.205	Likely Benign	0.660	Likely Pathogenic	Likely Benign	0.114	Likely Benign										1.31	Neutral	0.000	Benign	0.001	Benign	4.37	Benign	0.91	Tolerated	3.77	5	2	-3	-3.6	-30.03
c.2735C>A	T912N	Likely Benign		Uncertain		1				-4.260	Likely Benign	0.190	Likely Benign	Likely Benign	0.116	Likely Benign										-1.15	Neutral	0.999	Probably Damaging	0.977	Probably Damaging	3.96	Benign	0.00	Affected	3.77	5	0	0	-2.8	13.00
c.3308G>A	R1103H	Likely Benign		Benign/Likely benign		3	6-33443860-G-A	31	2.03e-5	-3.622	Likely Benign	0.156	Likely Benign	Likely Benign	0.116	Likely Benign										-1.97	Neutral	0.996	Probably Damaging	0.733	Possibly Damaging	2.49	Pathogenic	0.01	Affected	3.77	5	2	0	1.3	-19.05
c.1345A>G	S449G (3D Viewer)	Likely Benign	GAP	Uncertain		1	6-33438250-A-G	3	1.86e-6	-5.936	Likely Benign	0.071	Likely Benign	Likely Benign	0.116	Likely Benign	0.47	Likely Benign	0.0	0.55	Ambiguous	0.51	Ambiguous	0.85	Ambiguous	-2.32	Neutral	0.948	Possibly Damaging	0.124	Benign	3.35	Benign	0.13	Tolerated	3.37	32	0	1	0.4	-30.03
c.899C>T	S300F (3D Viewer)	Likely Pathogenic	C2	Uncertain		1				-10.222	Likely Pathogenic	0.353	Ambiguous	Likely Benign	0.117	Likely Benign	-0.29	Likely Benign	0.4	0.16	Likely Benign	-0.07	Likely Benign	0.04	Likely Benign	-2.66	Deleterious	0.975	Probably Damaging	0.596	Possibly Damaging	1.52	Pathogenic	0.01	Affected	3.47	19	-3	-2	3.6	60.10	233.6	-67.6	-0.1	0.0	0.4	0.2	X	X						Potentially Pathogenic	The hydroxyl group of the Ser300 side chain, located in a β hairpin loop linking two anti-parallel β sheet strands (res. Met289-Pro298, res. Thr305-Asn315), hydrogen bonds with the guanidinium group of Arg299 and the backbone amide group and side chain of Ser302. Thus, in the WT simulations, it contributes to the β hairpin stability. In the variant simulations, the phenol ring of Phe300 cannot form any side chain-related hydrogen bonds, and Arg299 is moved away from its central hairpin loop position.β hairpins are potential nucleation sites during the initial stages of protein folding, so even minor changes in them could be significant. Due to its location near the membrane surface, the residue swap could also affect the C2 loop dynamics and SynGAP-membrane association. However, this is beyond the scope of the solvent-only simulations to unravel.
c.3824G>T	R1275L			Likely Benign		1	6-33447872-G-T	1	6.45e-7	-6.052	Likely Benign	0.446	Ambiguous	Likely Benign	0.117	Likely Benign										-4.04	Deleterious	0.800	Possibly Damaging	0.277	Benign	2.55	Benign	0.01	Affected	3.77	5	-3	-2	8.3	-43.03
c.3977C>T	P1326L	Likely Benign		Uncertain		1				-5.541	Likely Benign	0.115	Likely Benign	Likely Benign	0.117	Likely Benign										-1.06	Neutral	0.999	Probably Damaging	0.994	Probably Damaging	3.62	Benign	0.00	Affected	3.77	5	-3	-3	5.4	16.04
c.379C>T	R127W			Uncertain		1				-4.776	Likely Benign	0.806	Likely Pathogenic	Ambiguous	0.118	Likely Benign										-2.98	Deleterious	0.989	Probably Damaging	0.420	Benign	3.88	Benign	0.00	Affected			2	-3	3.6	30.03
c.1222A>G	T408A		C2	Uncertain		1				-8.304	Likely Pathogenic	0.114	Likely Benign	Likely Benign	0.118	Likely Benign	0.37	Likely Benign	0.6	-0.06	Likely Benign	0.16	Likely Benign	0.72	Ambiguous	-3.07	Deleterious	0.540	Possibly Damaging	0.131	Benign	4.16	Benign	0.14	Tolerated			1	0	2.5	-30.03
c.431C>T	T144M	Likely Pathogenic		Uncertain		2	6-33432728-C-T	2	1.30e-6	-11.228	Likely Pathogenic	0.922	Likely Pathogenic	Ambiguous	0.118	Likely Benign										-3.16	Deleterious	0.913	Possibly Damaging	0.333	Benign	3.73	Benign	0.00	Affected	3.61	5	-1	-1	2.6	30.09
c.3304G>C	A1102P	Likely Benign		Uncertain		1				-5.120	Likely Benign	0.077	Likely Benign	Likely Benign	0.118	Likely Benign										-0.97	Neutral	0.000	Benign	0.002	Benign	2.26	Pathogenic	0.13	Tolerated	3.77	5	-1	1	-3.4	26.04
c.2684G>A	S895N	Likely Benign		Uncertain		1				-6.399	Likely Benign	0.604	Likely Pathogenic	Likely Benign	0.118	Likely Benign										-0.85	Neutral	0.991	Probably Damaging	0.988	Probably Damaging	2.64	Benign	0.30	Tolerated	4.32	4	1	1	-2.7	27.03
c.2753C>T	A918V	Likely Benign		Uncertain		3	6-33443305-C-T	2	1.24e-6	-3.684	Likely Benign	0.112	Likely Benign	Likely Benign	0.119	Likely Benign										-1.61	Neutral	0.980	Probably Damaging	0.782	Possibly Damaging	2.61	Benign	0.03	Affected	4.32	4	0	0	2.4	28.05
c.2521G>A	V841M			Uncertain		1	6-33443073-G-A	3	1.86e-6	-7.000	In-Between	0.651	Likely Pathogenic	Likely Benign	0.119	Likely Benign										-0.74	Neutral	0.999	Probably Damaging	0.998	Probably Damaging	2.54	Benign	0.02	Affected	3.77	5	1	2	-2.3	32.06
c.266C>T	P89L			Uncertain		2				-6.775	Likely Benign	0.982	Likely Pathogenic	Likely Pathogenic	0.119	Likely Benign										-3.29	Deleterious	0.889	Possibly Damaging	0.058	Benign	3.73	Benign	0.00	Affected	4.32	1	-3	-3	5.4	16.04
c.373C>T	P125S	Likely Benign		Uncertain		1				-3.769	Likely Benign	0.238	Likely Benign	Likely Benign	0.121	Likely Benign										-3.57	Deleterious	0.580	Possibly Damaging	0.140	Benign	2.86	Benign	0.02	Affected	3.61	5	1	-1	0.8	-10.04
c.3056G>A	R1019H	Likely Benign		Conflicting		2	6-33443608-G-A	67	4.15e-5	-4.610	Likely Benign	0.258	Likely Benign	Likely Benign	0.122	Likely Benign										-1.95	Neutral	0.995	Probably Damaging	0.845	Possibly Damaging	2.39	Pathogenic	0.01	Affected	3.77	5	2	0	1.3	-19.05
c.2900G>T	R967L	Likely Benign		Uncertain		1	6-33443452-G-T	1	6.20e-7	-3.496	Likely Benign	0.164	Likely Benign	Likely Benign	0.123	Likely Benign										-0.99	Neutral	0.959	Probably Damaging	0.586	Possibly Damaging	4.15	Benign	0.75	Tolerated	4.32	2	-2	-3	8.3	-43.03
c.3237C>A	S1079R	Likely Benign		Uncertain		1	6-33443789-C-A	4	2.51e-6	-4.579	Likely Benign	0.955	Likely Pathogenic	Ambiguous	0.123	Likely Benign										-1.81	Neutral	0.177	Benign	0.075	Benign	3.86	Benign	0.00	Affected	3.77	5	0	-1	-3.7	69.11
c.3932T>C	L1311P	Likely Benign		Likely Benign		1	6-33451806-T-C	1	6.21e-7	-1.831	Likely Benign	0.079	Likely Benign	Likely Benign	0.123	Likely Benign										-0.52	Neutral	0.579	Possibly Damaging	0.335	Benign	2.72	Benign	0.18	Tolerated	3.77	5	-3	-3	-5.4	-16.04
c.3237C>G	S1079R	Likely Benign		Benign		1				-4.579	Likely Benign	0.955	Likely Pathogenic	Ambiguous	0.124	Likely Benign										-1.81	Neutral	0.177	Benign	0.075	Benign	3.86	Benign	0.00	Affected	3.77	5	0	-1	-3.7	69.11
c.2353C>T	R785C	Likely Pathogenic	SH3-binding motif	Uncertain		1	6-33442905-C-T	29	1.80e-5	-5.887	Likely Benign	0.662	Likely Pathogenic	Likely Benign	0.126	Likely Benign										-5.06	Deleterious	0.144	Benign	0.046	Benign	2.22	Pathogenic	0.00	Affected	3.64	6	-4	-3	7.0	-53.05
c.4000A>G	N1334D			Uncertain		1	6-33451874-A-G			-4.584	Likely Benign	0.674	Likely Pathogenic	Likely Benign	0.126	Likely Benign										-3.06	Deleterious	0.886	Possibly Damaging	0.522	Possibly Damaging	3.55	Benign	0.00	Affected	3.77	5	1	2	0.0	0.98
c.2015C>T	T672M (3D Viewer)		GAP	Conflicting		2	6-33441274-C-T	19	1.18e-5	-9.472	Likely Pathogenic	0.174	Likely Benign	Likely Benign	0.127	Likely Benign	0.31	Likely Benign	0.4	1.52	Ambiguous	0.92	Ambiguous	0.41	Likely Benign	-4.34	Deleterious	0.993	Probably Damaging	0.520	Possibly Damaging	3.39	Benign	0.00	Affected	3.40	25	-1	-1	2.6	30.09	231.9	-52.9	1.1	0.1	0.5	0.0		X				X		Potentially Pathogenic	The hydroxyl group of Thr672, located in an entangled α-α loop connecting the two α-helices (res. Ser641-Glu666 and res. Leu685-Val699), is involved in a highly coordinated hydrogen-bonding network between residues from two α-helices (res. Ser641-Glu666 and res. Arg563-Glu578) and from the α-α loop itself, such as Lys566, Glu666, and Asn669. Met672 can only form a hydrogen bond with the amino group of the Lys566 side chain via its backbone carbonyl group. Nevertheless, the Lys566-Glu666 salt bridge forms intermittently. This is possible because Asn669 keeps the carboxylate group of Glu666 in the vicinity through hydrogen bonding, and the hydrophobic side chain of Met stays mostly rotated away from the salt bridge. Consequently, no drastic disruption of the hydrogen-bond network that keeps the loop close to the helices occurs in the variant simulations.
c.3956C>G	A1319G	Likely Benign		Uncertain		2	6-33451830-C-G			-3.927	Likely Benign	0.084	Likely Benign	Likely Benign	0.128	Likely Benign										-0.74	Neutral	0.819	Possibly Damaging	0.581	Possibly Damaging	4.07	Benign	0.06	Tolerated	3.77	5	1	0	-2.2	-14.03
c.3022G>A	D1008N	Likely Benign		Likely Benign		1	6-33443574-G-A	3	1.86e-6	-4.045	Likely Benign	0.714	Likely Pathogenic	Likely Benign	0.128	Likely Benign										-2.15	Neutral	0.999	Probably Damaging	0.997	Probably Damaging	2.75	Benign	0.01	Affected	3.77	5	2	1	0.0	-0.98
c.2354G>A	R785H		SH3-binding motif	Uncertain		2	6-33442906-G-A	4	2.50e-6	-4.782	Likely Benign	0.388	Ambiguous	Likely Benign	0.129	Likely Benign										-2.61	Deleterious	0.999	Probably Damaging	0.947	Probably Damaging	2.25	Pathogenic	0.01	Affected	3.64	6	2	0	1.3	-19.05
c.3290C>T	P1097L	Likely Benign		Benign		1				-4.410	Likely Benign	0.145	Likely Benign	Likely Benign	0.131	Likely Benign										-2.07	Neutral	0.611	Possibly Damaging	0.198	Benign	2.64	Benign	0.05	Affected	3.77	5	-3	-3	5.4	16.04
c.558G>C	L186F	Likely Pathogenic		Uncertain		1				-11.861	Likely Pathogenic	0.993	Likely Pathogenic	Likely Pathogenic	0.132	Likely Benign										-3.03	Deleterious	0.009	Benign	0.012	Benign	3.50	Benign	0.00	Affected			2	0	-1.0	34.02
c.1004G>A	R335H (3D Viewer)	Likely Pathogenic	C2	Uncertain		1	6-33437909-G-A	2	1.24e-6	-12.521	Likely Pathogenic	0.831	Likely Pathogenic	Ambiguous	0.132	Likely Benign	0.58	Ambiguous	0.1	0.22	Likely Benign	0.40	Likely Benign	0.72	Ambiguous	-3.02	Deleterious	1.000	Probably Damaging	0.998	Probably Damaging	1.70	Pathogenic	0.03	Affected	3.38	22	2	0	1.3	-19.05	242.4	82.1	-2.4	0.6	-0.1	0.1								Uncertain	The guanidinium group of Arg335, located in a β hairpin loop linking two anti-parallel β sheet strands (res. Ala322-Asp330, res. Gly341-Pro349), faces the post-synaptic inner membrane surface. In the WT simulations, the Arg335 side chain dynamically forms salt bridges with the carboxylate groups of Asp322, Asp338, and Asp616. In contrast, the imidazole ring of His335, which is not double protonated and thus not positively charged in the variant simulations, continues to move dynamically without forming any lasting or strong interactions. Importantly, the positively charged arginine residues of the C2 domain are ideal membrane anchors for ensuring SynGAP-membrane association. However, this phenomenon cannot be addressed using solvent-only simulations.
c.2101C>T	P701S (3D Viewer)	Likely Benign	GAP	Uncertain		1	6-33441360-C-T	3	1.86e-6	-4.375	Likely Benign	0.221	Likely Benign	Likely Benign	0.132	Likely Benign	1.33	Ambiguous	0.0	0.12	Likely Benign	0.73	Ambiguous	-0.36	Likely Benign	0.78	Neutral	0.044	Benign	0.025	Benign	3.48	Benign	1.00	Tolerated	3.47	10	-1	1	0.8	-10.04																10.1016/j.ajhg.2020.11.011
c.3977C>G	P1326R	Likely Benign		Uncertain		1				-5.097	Likely Benign	0.240	Likely Benign	Likely Benign	0.133	Likely Benign										-0.82	Neutral	0.999	Probably Damaging	0.994	Probably Damaging	3.62	Benign	0.00	Affected	3.77	5	0	-2	-2.9	59.07
c.3313C>T	R1105W			Uncertain		1	6-33443865-C-T	6	3.93e-6	-6.911	Likely Benign	0.488	Ambiguous	Likely Benign	0.133	Likely Benign										-4.34	Deleterious	0.999	Probably Damaging	0.696	Possibly Damaging	2.42	Pathogenic	0.02	Affected	3.77	5	-3	2	3.6	30.03
c.223G>A	E75K	Likely Benign		Benign/Likely benign		2				-4.020	Likely Benign	0.358	Ambiguous	Likely Benign	0.134	Likely Benign										-1.12	Neutral	0.748	Possibly Damaging	0.017	Benign	4.07	Benign	0.00	Affected			0	1	-0.4	-0.94
c.3223C>A	Q1075K	Likely Benign		Uncertain		1				-5.135	Likely Benign	0.728	Likely Pathogenic	Likely Benign	0.134	Likely Benign										-0.67	Neutral	0.963	Probably Damaging	0.959	Probably Damaging	2.75	Benign	1.00	Tolerated	3.77	5	1	1	-0.4	0.04
c.3251C>A	P1084H	Likely Benign		Uncertain		1	6-33443803-C-A	1	6.31e-7	-4.125	Likely Benign	0.323	Likely Benign	Likely Benign	0.134	Likely Benign										-3.16	Deleterious	0.997	Probably Damaging	0.840	Possibly Damaging	3.96	Benign	0.00	Affected	3.77	5	-2	0	-1.6	40.02
c.2369C>G	T790S	Likely Benign	SH3-binding motif	Uncertain		1				-3.914	Likely Benign	0.123	Likely Benign	Likely Benign	0.134	Likely Benign										-1.83	Neutral	0.997	Probably Damaging	0.989	Probably Damaging	2.39	Pathogenic	0.33	Tolerated	3.64	6	1	1	-0.1	-14.03
c.2962C>T	L988F	Likely Benign		Uncertain		1	6-33443514-C-T	1	6.20e-7	-4.368	Likely Benign	0.356	Ambiguous	Likely Benign	0.135	Likely Benign										-1.70	Neutral	0.977	Probably Damaging	0.900	Possibly Damaging	2.69	Benign	0.00	Affected	4.32	2	2	0	-1.0	34.02
c.2818G>A	G940S	Likely Benign		Uncertain		1	6-33443370-G-A	1	6.20e-7	-5.451	Likely Benign	0.084	Likely Benign	Likely Benign	0.135	Likely Benign										0.45	Neutral	0.409	Benign	0.253	Benign	2.77	Benign	0.44	Tolerated	3.77	5	1	0	-0.4	30.03
c.515G>A	R172Q			Uncertain		1	6-33435157-G-A	3	1.86e-6	-7.245	In-Between	0.465	Ambiguous	Likely Benign	0.135	Likely Benign										-1.72	Neutral	0.804	Possibly Damaging	0.091	Benign	4.04	Benign	0.04	Affected	3.61	5	1	1	1.0	-28.06
c.2948G>A	S983N			Likely Benign		1	6-33443500-G-A	6	3.72e-6	-5.604	Likely Benign	0.909	Likely Pathogenic	Ambiguous	0.136	Likely Benign										-1.78	Neutral	0.991	Probably Damaging	0.988	Probably Damaging	2.04	Pathogenic	0.00	Affected	4.32	1	1	1	-2.7	27.03
c.2300T>C	I767T	Likely Benign		Uncertain		1				-3.749	Likely Benign	0.252	Likely Benign	Likely Benign	0.138	Likely Benign										-0.78	Neutral	0.625	Possibly Damaging	0.249	Benign	4.12	Benign	0.46	Tolerated	3.64	6	0	-1	-5.2	-12.05
c.2443C>A	R815S		SH3-binding motif	Benign		1				-7.324	In-Between	0.950	Likely Pathogenic	Ambiguous	0.138	Likely Benign										-1.86	Neutral	0.999	Probably Damaging	0.997	Probably Damaging	2.67	Benign	0.02	Affected			0	-1	3.7	-69.11
c.3977C>A	P1326Q	Likely Benign		Uncertain		1	6-33451851-C-A	1	6.40e-7	-5.422	Likely Benign	0.128	Likely Benign	Likely Benign	0.138	Likely Benign										-0.86	Neutral	0.999	Probably Damaging	0.994	Probably Damaging	3.62	Benign	0.00	Affected	3.77	5	-1	0	-1.9	31.01
c.2909A>G	E970G	Likely Benign		Benign		1				-0.167	Likely Benign	0.139	Likely Benign	Likely Benign	0.139	Likely Benign										-0.93	Neutral	0.144	Benign	0.058	Benign	4.09	Benign	0.10	Tolerated	4.32	2	0	-2	3.1	-72.06
c.1964T>A	L655Q (3D Viewer)	Likely Benign	GAP	Uncertain		1				-5.278	Likely Benign	0.144	Likely Benign	Likely Benign	0.139	Likely Benign	-0.01	Likely Benign	0.0	0.69	Ambiguous	0.34	Likely Benign	-0.15	Likely Benign	0.61	Neutral	0.955	Possibly Damaging	0.602	Possibly Damaging	3.59	Benign	0.65	Tolerated	3.39	24	-2	-2	-7.3	14.97	229.9	-8.6	0.0	0.0	0.4	0.0						X		Potentially Benign	The iso-butyl side chain of Leu655, located on the surface of an α helix (res. Ser641-Glu666), is not involved in any interactions in the WT simulations. In the variant simulations, the carboxamide side chain of Gln655 dynamically interacts with neighboring residues (e.g., Glu651, Glu656, Arg544) on the protein surface, with no negative structural effects.
c.250C>G	R84G			Uncertain		1				-6.627	Likely Benign	0.989	Likely Pathogenic	Likely Pathogenic	0.139	Likely Benign										-2.64	Deleterious	0.962	Probably Damaging	0.726	Possibly Damaging	3.68	Benign	0.00	Affected	4.32	1	-3	-2	4.1	-99.14
c.3307C>T	R1103C			Uncertain		1	6-33443859-C-T	6	3.92e-6	-2.440	Likely Benign	0.246	Likely Benign	Likely Benign	0.140	Likely Benign										-3.01	Deleterious	0.996	Probably Damaging	0.787	Possibly Damaging	2.41	Pathogenic	0.01	Affected	3.77	5	-3	-4	7.0	-53.05
c.113C>T	P38L	Likely Benign		Conflicting		4	6-33423522-C-T	8	4.96e-6	-2.469	Likely Benign	0.197	Likely Benign	Likely Benign	0.141	Likely Benign										-2.56	Deleterious	0.983	Probably Damaging	0.931	Probably Damaging	4.02	Benign	0.00	Affected	4.32	1	-3	-3	5.4	16.04
c.2812G>A	G938R	Likely Benign		Uncertain		1				-5.271	Likely Benign	0.732	Likely Pathogenic	Likely Benign	0.141	Likely Benign										-1.11	Neutral	0.999	Probably Damaging	0.985	Probably Damaging	2.74	Benign	0.36	Tolerated	3.77	5	-3	-2	-4.1	99.14
c.3009C>G	S1003R			Uncertain		1				-5.113	Likely Benign	0.991	Likely Pathogenic	Likely Pathogenic	0.141	Likely Benign										-1.88	Neutral	0.999	Probably Damaging	0.996	Probably Damaging	2.48	Pathogenic	0.00	Affected	3.77	5	0	-1	-3.7	69.11
c.453C>A	D151E	Likely Benign		Uncertain		1				-5.662	Likely Benign	0.886	Likely Pathogenic	Ambiguous	0.142	Likely Benign										-2.02	Neutral	0.984	Probably Damaging	0.967	Probably Damaging	3.99	Benign	0.11	Tolerated	3.61	5	3	2	0.0	14.03
c.3980C>T	P1327L	Likely Benign		Uncertain		1	6-33451854-C-T	2	1.28e-6	-5.264	Likely Benign	0.242	Likely Benign	Likely Benign	0.142	Likely Benign										-1.24	Neutral	0.994	Probably Damaging	0.908	Possibly Damaging	4.12	Benign	0.10	Tolerated	3.77	5	-3	-3	5.4	16.04
c.2113A>C	K705Q (3D Viewer)	Likely Benign	GAP	Uncertain		1	6-33441372-A-C	1	6.20e-7	-5.787	Likely Benign	0.436	Ambiguous	Likely Benign	0.142	Likely Benign	-0.10	Likely Benign	0.0	0.33	Likely Benign	0.12	Likely Benign	-0.02	Likely Benign	-0.24	Neutral	0.997	Probably Damaging	0.969	Probably Damaging	3.42	Benign	0.78	Tolerated	3.47	10	1	1	0.4	-0.04
c.2750C>G	P917R	Likely Benign		Uncertain		1	6-33443302-C-G	5	3.10e-6	-4.475	Likely Benign	0.363	Ambiguous	Likely Benign	0.142	Likely Benign										-1.70	Neutral	0.642	Possibly Damaging	0.316	Benign	2.68	Benign	0.00	Affected	3.77	5	-2	0	-2.9	59.07
c.2458T>A	Y820N			Uncertain		1				-9.032	Likely Pathogenic	0.842	Likely Pathogenic	Ambiguous	0.143	Likely Benign										-1.53	Neutral	0.999	Probably Damaging	0.977	Probably Damaging	2.74	Benign	0.20	Tolerated			-2	-2	-2.2	-49.07
c.2860C>T	P954S	Likely Benign		Likely Benign		1	6-33443412-C-T	1	6.20e-7	-3.525	Likely Benign	0.062	Likely Benign	Likely Benign	0.143	Likely Benign										-0.25	Neutral	0.954	Possibly Damaging	0.812	Possibly Damaging	2.87	Benign	1.00	Tolerated	3.77	5	1	-1	0.8	-10.04
c.1888A>G	I630V (3D Viewer)		GAP	Benign/Likely benign		4	6-33440940-A-G	59	3.66e-5	-7.264	In-Between	0.145	Likely Benign	Likely Benign	0.143	Likely Benign	1.33	Ambiguous	0.0	0.94	Ambiguous	1.14	Ambiguous	0.64	Ambiguous	-0.38	Neutral	0.018	Benign	0.011	Benign	-1.37	Pathogenic	0.35	Tolerated	3.37	34	4	3	-0.3	-14.03	235.0	26.2	-0.1	0.0	-0.3	0.1						X		Potentially Benign	The sec-butyl side chain of Ile630, located in an α helix (res. Glu617-Asn635), packs with hydrophobic residues (e.g., Phe594, Leu633, Ile626, Ile602) in the hydrophobic inter-helix space between two α helices (res. Glu617-Asn635 and res. Glu582-Met603).In the variant simulations, the iso-propyl side chain of Val630, which shares a similar size and physicochemical properties with Ile630 in the WT, maintains similar interactions in the inter-helix space. Although no negative structural effects are observed during the simulations, the implications of the residue swap on the complex formation with the GTPase, due to its location, cannot be investigated using solvent-only simulations.
c.3640C>T	R1214W	Likely Pathogenic	Coiled-coil	Uncertain		1	6-33446632-C-T	2	1.24e-6	-8.799	Likely Pathogenic	0.710	Likely Pathogenic	Likely Benign	0.143	Likely Benign										-4.95	Deleterious	1.000	Probably Damaging	0.983	Probably Damaging	2.45	Pathogenic	0.00	Affected	3.77	5	2	-3	3.6	30.03
c.913A>G	T305A (3D Viewer)	Likely Benign	C2	Conflicting		2	6-33437818-A-G	13	8.05e-6	-4.307	Likely Benign	0.078	Likely Benign	Likely Benign	0.144	Likely Benign	1.30	Ambiguous	0.6	1.55	Ambiguous	1.43	Ambiguous	0.77	Ambiguous	-2.10	Neutral	0.939	Possibly Damaging	0.645	Possibly Damaging	1.76	Pathogenic	0.12	Tolerated	3.40	20	1	0	2.5	-30.03	177.9	43.5	-0.2	0.1	0.4	0.0								Uncertain	The hydroxyl group of Thr305, located at the beginning of an anti-parallel β strand (res. Thr305-Asn315), hydrogen bonds with the carboxylate groups of Glu270 and Asp304 in the anti-parallel β strand and the adjacent β hairpin loop, respectively. In the variant simulations, the methyl group of the Ala305 side chain cannot hydrogen bond with either of the acidic residues, which could weaken the integrity of the tertiary structure and the β hairpin loop. Indeed, the guanidinium group of Arg299 does not acquire its central hairpin loop position due to the residue swap.β hairpins are potential nucleation sites during the initial stages of protein folding, so even minor changes in them could be significant. Due to its location near the membrane surface, the residue swap could also affect the C2 loop dynamics and SynGAP-membrane association. However, this is beyond the scope of the solvent-only simulations to unravel.
c.1973G>A	G658D (3D Viewer)		GAP	Uncertain		1	6-33441232-G-A	3	1.86e-6	-7.786	In-Between	0.442	Ambiguous	Likely Benign	0.144	Likely Benign	-0.40	Likely Benign	0.1	-0.59	Ambiguous	-0.50	Ambiguous	0.46	Likely Benign	-2.64	Deleterious	0.008	Benign	0.005	Benign	3.53	Benign	0.38	Tolerated	3.39	24	1	-1	-3.1	58.04	219.8	-84.3	0.0	0.0	0.2	0.1						X		Potentially Pathogenic	Gly658, located on the outer surface of an α helix (res. Ser641-Glu666), weakens the helix integrity at that spot, which is necessary for the kink in the middle of the long helix. In the variant simulations, the carboxylic acid side chain of Asp658 is on the surface of the α helix and is not involved in any interactions. However, aspartate is not as effective a breaker of the secondary structure element as glycine, which may lead to misfolding.
c.3238G>A	A1080T	Likely Benign		Conflicting		2	6-33443790-G-A	17	1.06e-5	-3.928	Likely Benign	0.133	Likely Benign	Likely Benign	0.144	Likely Benign										-0.19	Neutral	0.253	Benign	0.042	Benign	4.10	Benign	0.60	Tolerated	3.77	5	1	0	-2.5	30.03
c.2582C>T	S861L	Likely Benign		Uncertain		1	6-33443134-C-T	2	1.24e-6	-4.966	Likely Benign	0.219	Likely Benign	Likely Benign	0.144	Likely Benign										-2.10	Neutral	0.904	Possibly Damaging	0.355	Benign	3.93	Benign	0.07	Tolerated	4.32	3	-3	-2	4.6	26.08
c.3323G>T	S1108I			Uncertain		1	6-33443875-G-T			-3.666	Likely Benign	0.292	Likely Benign	Likely Benign	0.145	Likely Benign										-3.73	Deleterious	0.971	Probably Damaging	0.604	Possibly Damaging	2.44	Pathogenic	0.10	Tolerated	3.77	5	-2	-1	5.3	26.08
c.3196C>T	P1066S	Likely Benign		Likely Pathogenic		1				-4.746	Likely Benign	0.070	Likely Benign	Likely Benign	0.145	Likely Benign										-2.47	Neutral	0.972	Probably Damaging	0.850	Possibly Damaging	2.74	Benign	0.00	Affected	4.32	2	1	-1	0.8	-10.04
c.59C>G	P20R	Likely Benign		Uncertain		1				-3.548	Likely Benign	0.434	Ambiguous	Likely Benign	0.146	Likely Benign										-0.15	Neutral	0.972	Probably Damaging	0.804	Possibly Damaging	4.33	Benign	0.00	Affected	4.32	1	0	-2	-2.9	59.07
c.2443C>G	R815G		SH3-binding motif	Uncertain		1				-7.983	In-Between	0.854	Likely Pathogenic	Ambiguous	0.146	Likely Benign										-3.22	Deleterious	0.999	Probably Damaging	0.997	Probably Damaging	2.62	Benign	0.02	Affected	4.32	4	-3	-2	4.1	-99.14
c.43G>C	A15P	Likely Benign		Uncertain		1				-3.436	Likely Benign	0.097	Likely Benign	Likely Benign	0.146	Likely Benign										-0.23	Neutral	0.880	Possibly Damaging	0.123	Benign	4.09	Benign	0.00	Affected			1	-1	-3.4	26.04
c.3974C>T	P1325L	Likely Benign		Uncertain		1	6-33451848-C-T			-5.256	Likely Benign	0.085	Likely Benign	Likely Benign	0.146	Likely Benign										-1.05	Neutral	0.000	Benign	0.000	Benign	4.05	Benign	0.00	Affected	4.32	1	-3	-3	5.4	16.04
c.1957C>G	L653V	Likely Benign	GAP	Uncertain		1				-7.050	In-Between	0.301	Likely Benign	Likely Benign	0.146	Likely Benign	3.28	Destabilizing	0.3	2.18	Destabilizing	2.73	Destabilizing	1.32	Destabilizing	-2.25	Neutral	0.227	Benign	0.039	Benign	3.28	Benign	0.08	Tolerated			2	1	0.4	-14.03
c.2249G>A	G750E			Uncertain		1				-2.618	Likely Benign	0.413	Ambiguous	Likely Benign	0.146	Likely Benign										-2.27	Neutral	1.000	Probably Damaging	0.982	Probably Damaging	2.49	Pathogenic	0.01	Affected	3.99	5	0	-2	-3.1	72.06
c.1260T>G	F420L (3D Viewer)	Likely Pathogenic	GAP	Uncertain		1				-8.432	Likely Pathogenic	0.998	Likely Pathogenic	Likely Pathogenic	0.146	Likely Benign	1.76	Ambiguous	0.0	1.41	Ambiguous	1.59	Ambiguous	1.04	Destabilizing	-5.39	Deleterious	0.009	Benign	0.005	Benign	4.22	Benign	0.39	Tolerated	3.37	29	2	0	1.0	-34.02	231.1	13.2	0.0	0.0	-0.1	0.0						X		Potentially Benign	In the WT, the phenyl ring of the Phe420 side chain, located on an α helix (res. Met414-Glu436), packs against hydrophobic residues in the interhelix area of the GAP domain (e.g., Leu689, Leu714, Leu717, Leu718). In the variant simulations, the iso-butyl side chain of Leu420 also packs into the hydrophobic inter-helix niche, but due to its smaller size, the resulting steric interactions are not as favorable as with phenylalanine. In short, the residue swap does not cause severe effects on the protein structure based on the variant simulations.
c.303C>A	H101Q	Likely Benign		Uncertain		1	6-33432168-C-A	1	6.20e-7	-2.827	Likely Benign	0.124	Likely Benign	Likely Benign	0.147	Likely Benign										-0.37	Neutral	0.824	Possibly Damaging	0.880	Possibly Damaging	4.24	Benign	0.00	Affected	4.32	1	3	0	-0.3	-9.01
c.611C>G	S204C (3D Viewer)	Likely Benign	PH	Uncertain		1				-6.613	Likely Benign	0.127	Likely Benign	Likely Benign	0.148	Likely Benign	0.65	Ambiguous	0.4	-1.13	Ambiguous	-0.24	Likely Benign	0.10	Likely Benign	-0.64	Neutral	0.978	Probably Damaging	0.753	Possibly Damaging	4.13	Benign	0.05	Affected	3.44	10	0	-1	3.3	16.06	223.6	-13.8	0.6	0.3	0.0	0.2	X							Uncertain	The hydroxyl-containing Ser204, located in the N-terminal loop before the first anti-parallel β sheet strand (res. Ile205-Pro208), is replaced by the thiol-containing cysteine. In the WT simulations, Ser204 simultaneously forms hydrogen bonds with the backbone carbonyl of Asp201 and the hydroxyl group of Thr224, helping to stabilize the two anti-parallel β strands (res. Ile205-Lys207 and Cys219-Thr223) at the end of the β sheet. Since the thiol group of cysteine forms weaker hydrogen bonds than the hydroxyl group of serine, Cys204 does not maintain the hydrogen bond network as stably as Ser204 in the variant simulations. However, because the model ends abruptly at the N-terminus, no definite conclusions can be drawn from the simulations.
c.3821G>A	R1274H			Likely Benign		1	6-33447869-G-A	4	2.58e-6	-5.259	Likely Benign	0.256	Likely Benign	Likely Benign	0.149	Likely Benign										-3.20	Deleterious	1.000	Probably Damaging	0.995	Probably Damaging	2.49	Pathogenic	0.01	Affected	3.77	5	0	2	1.3	-19.05
c.2809G>C	D937H	Likely Benign		Uncertain		1				-0.733	Likely Benign	0.677	Likely Pathogenic	Likely Benign	0.150	Likely Benign										-1.74	Neutral	1.000	Probably Damaging	0.975	Probably Damaging	2.68	Benign	0.13	Tolerated	3.77	5	-1	1	0.3	22.05
c.484C>T	R162C			Pathogenic		2				-8.157	Likely Pathogenic	0.787	Likely Pathogenic	Ambiguous	0.150	Likely Benign										-2.05	Neutral	0.988	Probably Damaging	0.513	Possibly Damaging	4.00	Benign	0.11	Tolerated	3.74	4	-4	-3	7.0	-53.05
c.901G>A	A301T (3D Viewer)	Likely Benign	C2	Uncertain		5	6-33437806-G-A	2	1.24e-6	-3.448	Likely Benign	0.070	Likely Benign	Likely Benign	0.150	Likely Benign	0.36	Likely Benign	0.2	-0.33	Likely Benign	0.02	Likely Benign	0.03	Likely Benign	-0.25	Neutral	0.997	Probably Damaging	0.989	Probably Damaging	4.15	Benign	0.22	Tolerated	4.32	14	1	0	-2.5	30.03	219.8	-42.8	-0.1	0.0	-0.5	0.2								Uncertain	The methyl group of Ala301, located in a β hairpin loop linking two anti-parallel β sheet strands (res. Met289-Pro298, res. Thr305-Asn315), points outward from the β hairpin loop, and its backbone atoms do not participate in the loop formation in the WT simulations. In the variant simulations, the hydroxyl group of the Thr301 side chain also mostly points outward; however, the guanidinium group of Arg299 is moved away from its central hairpin loop position.β hairpins are potential nucleation sites during the initial stages of protein folding, so even minor changes in them could be significant. Due to its location near the membrane surface, the residue swap could also affect the C2 loop dynamics and SynGAP-membrane association. However, this is beyond the scope of the solvent-only simulations to unravel.
c.865A>G	M289V (3D Viewer)	Likely Benign	C2	Benign		1				-4.239	Likely Benign	0.117	Likely Benign	Likely Benign	0.150	Likely Benign	1.09	Ambiguous	0.1	-0.27	Likely Benign	0.41	Likely Benign	0.24	Likely Benign	-0.36	Neutral	0.136	Benign	0.054	Benign	1.80	Pathogenic	1.00	Tolerated	3.38	23	2	1	2.3	-32.06	204.2	51.0	0.0	0.0	0.2	0.0						X		Potentially Benign	The hydrophobic residue Met289, located in a β hairpin linking two anti-parallel β sheet strands (res. Met289-Arg299, res. Arg272-Leu286), is swapped for another hydrophobic residue, valine. In the variant simulations, the branched hydrocarbon side chain of Val289 packs against the phenol group of the Tyr291 side chain but is unable to form methionine-aromatic interactions. β hairpins are potential nucleation sites during the initial stages of protein folding, so even minor changes in them could be significant. However, based on the simulations, the residue swap does not cause adverse effects on the structure.
c.3326T>C	L1109P	Likely Benign		Conflicting		2				-5.313	Likely Benign	0.120	Likely Benign	Likely Benign	0.151	Likely Benign										-0.52	Neutral	0.002	Benign	0.003	Benign	2.65	Benign	0.07	Tolerated	4.32	2	-3	-3	-5.4	-16.04
c.2650C>T	R884W	Likely Benign		Uncertain		1	6-33443202-C-T	5	3.10e-6	-3.785	Likely Benign	0.332	Likely Benign	Likely Benign	0.151	Likely Benign										0.26	Neutral	0.995	Probably Damaging	0.812	Possibly Damaging	2.56	Benign	0.05	Affected	4.32	4	-3	2	3.6	30.03
c.2246G>A	R749Q	Likely Benign		Likely Benign		1	6-33441711-G-A	4	2.48e-6	-3.069	Likely Benign	0.212	Likely Benign	Likely Benign	0.152	Likely Benign										-1.00	Neutral	0.999	Probably Damaging	0.994	Probably Damaging	2.64	Benign	0.03	Affected	4.32	2	1	1	1.0	-28.06
c.583G>C	A195P	Likely Pathogenic		Likely Pathogenic		1				-9.715	Likely Pathogenic	0.978	Likely Pathogenic	Likely Pathogenic	0.152	Likely Benign										-3.03	Deleterious	0.997	Probably Damaging	0.916	Probably Damaging	4.00	Benign	0.04	Affected	3.54	6	1	-1	-3.4	26.04
c.2405G>A	G802D	Likely Benign	SH3-binding motif	Uncertain		1	6-33442957-G-A	1	6.20e-7	-5.083	Likely Benign	0.476	Ambiguous	Likely Benign	0.153	Likely Benign										-0.38	Neutral	0.126	Benign	0.138	Benign	2.72	Benign	0.09	Tolerated	3.77	5	1	-1	-3.1	58.04
c.2029A>T	S677C (3D Viewer)	Likely Benign	GAP	Benign		1				-8.496	Likely Pathogenic	0.076	Likely Benign	Likely Benign	0.153	Likely Benign	-0.51	Ambiguous	0.3	-0.30	Likely Benign	-0.41	Likely Benign	0.15	Likely Benign	-2.41	Neutral	0.932	Possibly Damaging	0.222	Benign	3.25	Benign	0.04	Affected	3.41	23	-1	0	3.3	16.06
c.2210A>C	Q737P	Likely Benign		Uncertain		1				-2.407	Likely Benign	0.054	Likely Benign	Likely Benign	0.154	Likely Benign										-1.22	Neutral	0.005	Benign	0.013	Benign	2.78	Benign	0.04	Affected	4.07	3	-1	0	1.9	-31.01
c.3731G>A	S1244N	Likely Pathogenic	Coiled-coil	Uncertain		1				-9.008	Likely Pathogenic	0.751	Likely Pathogenic	Likely Benign	0.154	Likely Benign										-1.87	Neutral	0.997	Probably Damaging	0.992	Probably Damaging	2.10	Pathogenic	0.15	Tolerated	3.77	5	1	1	-2.7	27.03
c.3907G>A	G1303S	Likely Benign		Uncertain		1				-2.271	Likely Benign	0.125	Likely Benign	Likely Benign	0.155	Likely Benign										-0.19	Neutral	0.649	Possibly Damaging	0.433	Benign	2.84	Benign	0.18	Tolerated			1	0	-0.4	30.03
c.986G>A	R329H (3D Viewer)	Likely Pathogenic	C2	Uncertain		1	6-33437891-G-A	2	1.24e-6	-10.154	Likely Pathogenic	0.769	Likely Pathogenic	Likely Benign	0.155	Likely Benign	2.53	Destabilizing	0.7	0.71	Ambiguous	1.62	Ambiguous	0.82	Ambiguous	-3.17	Deleterious	0.995	Probably Damaging	0.778	Possibly Damaging	4.04	Benign	0.05	Affected	3.41	15	2	0	1.3	-19.05	220.4	81.4	0.1	0.1	0.2	0.3								Uncertain	The guanidinium group of Arg329, located at the end of an anti-parallel β sheet strand (res. Ala322-Asp330), faces the negatively charged lipid bilayer surface. While the residue swap does not cause any apparent negative effects on the protein structure in the variant simulations, it could adversely affect the SynGAP-membrane association in reality. The positively charged Arg329 side chain forms hydrogen bonds with other loop residues (e.g., Ser371, Asp338) that are expected to dynamically interact with the membrane head group region. However, this phenomenon is beyond the scope of the solvent-only simulations to unravel. Notably, histidine can also be double protonated and positively charged, but this alternative protonation state was not considered in the variant simulations.
c.2840G>C	G947A	Likely Benign		Likely Benign		1	6-33443392-G-C	28	1.73e-5	-6.511	Likely Benign	0.080	Likely Benign	Likely Benign	0.156	Likely Benign										-0.41	Neutral	0.224	Benign	0.131	Benign	4.97	Benign	0.10	Tolerated	4.32	4	1	0	2.2	14.03
c.1286G>A	R429Q (3D Viewer)	Likely Benign	GAP	Uncertain		2	6-33438191-G-A	10	6.20e-6	-8.227	Likely Pathogenic	0.143	Likely Benign	Likely Benign	0.156	Likely Benign	0.45	Likely Benign	0.1	0.36	Likely Benign	0.41	Likely Benign	0.98	Ambiguous	-1.25	Neutral	1.000	Probably Damaging	0.979	Probably Damaging	3.47	Benign	0.58	Tolerated	3.38	25	1	1	1.0	-28.06	235.8	59.5	0.0	0.0	-0.3	0.4		X						Potentially Pathogenic	The guanidinium group of the Arg429 side chain, located in an α helix (res. Met414-Glu436), either forms a salt bridge with the carboxylate group of an acidic residue (Asp474, Asp467) or an H-bond with the hydroxyl group of Ser471 in an opposing α helix (res. Ala461-Phe476). In the variant simulations, Gln429 cannot form ionic interactions with the acidic residues; however, the carboxamide group can form multiple H-bonds. The H-bonding coordination of the Asn429 side chain varied between the replica simulations. In one simulation, three H-bonds were formed simultaneously with the Asp467 side chain, the backbone carbonyl group of Asn426, and the amide group of Met430 at the end of the same α helix. The residue swap could affect the tertiary structure assembly during folding due to weaker bond formation, but no large-scale negative effects were seen during the simulations.
c.3049T>C	F1017L	Likely Benign		Benign		1				-2.048	Likely Benign	0.934	Likely Pathogenic	Ambiguous	0.157	Likely Benign										-2.38	Neutral	0.798	Possibly Damaging	0.373	Benign	2.65	Benign	0.72	Tolerated	3.77	5	0	2	1.0	-34.02
c.2444G>A	R815H		SH3-binding motif	Likely Benign		2	6-33442996-G-A	24	1.49e-5	-7.474	In-Between	0.553	Ambiguous	Likely Benign	0.157	Likely Benign										-1.81	Neutral	1.000	Probably Damaging	0.998	Probably Damaging	2.61	Benign	0.02	Affected	4.32	4	2	0	1.3	-19.05																10.1016/j.ajhg.2020.11.011
c.3233T>A	V1078D	Likely Benign		Uncertain		1				-5.155	Likely Benign	0.979	Likely Pathogenic	Likely Pathogenic	0.158	Likely Benign										-1.45	Neutral	0.003	Benign	0.008	Benign	3.84	Benign	0.00	Affected	3.77	5	-3	-2	-7.7	15.96
c.73C>T	R25W	Likely Benign		Uncertain		2	6-33423482-C-T	6	3.72e-6	-5.133	Likely Benign	0.549	Ambiguous	Likely Benign	0.158	Likely Benign										-1.60	Neutral	0.994	Probably Damaging	0.919	Probably Damaging	3.92	Benign	0.00	Affected	4.32	1	-3	2	3.6	30.03
c.172A>G	M58V	Likely Benign		Uncertain		1				-2.211	Likely Benign	0.688	Likely Pathogenic	Likely Benign	0.160	Likely Benign										-0.71	Neutral	0.006	Benign	0.091	Benign	4.19	Benign	0.00	Affected	4.32	1	1	2	2.3	-32.06
c.3160G>A	G1054S	Likely Benign		Benign		1	6-33443712-G-A	32	1.99e-5	-5.294	Likely Benign	0.075	Likely Benign	Likely Benign	0.160	Likely Benign										0.21	Neutral	0.121	Benign	0.013	Benign	4.04	Benign	0.63	Tolerated	3.77	5	1	0	-0.4	30.03
c.458C>A	T153N	Likely Benign		Conflicting		3				-0.739	Likely Benign	0.226	Likely Benign	Likely Benign	0.161	Likely Benign										0.88	Neutral	0.888	Possibly Damaging	0.537	Possibly Damaging	4.23	Benign	0.81	Tolerated	3.61	5	0	0	-2.8	13.00
c.2434C>T	P812S	Likely Benign	SH3-binding motif	Uncertain		1	6-33442986-C-T	1	6.20e-7	-5.689	Likely Benign	0.456	Ambiguous	Likely Benign	0.162	Likely Benign										-0.62	Neutral	0.999	Probably Damaging	0.966	Probably Damaging	2.89	Benign	0.95	Tolerated	4.32	4	1	-1	0.8	-10.04
c.3262A>G	S1088G	Likely Benign		Uncertain		1				-5.034	Likely Benign	0.285	Likely Benign	Likely Benign	0.163	Likely Benign										-1.83	Neutral	0.979	Probably Damaging	0.973	Probably Damaging	2.63	Benign	0.03	Affected	3.77	5	0	1	0.4	-30.03
c.3287A>C	E1096A	Likely Benign		Uncertain		1				-4.504	Likely Benign	0.510	Ambiguous	Likely Benign	0.164	Likely Benign										-1.37	Neutral	0.626	Possibly Damaging	0.184	Benign	2.77	Benign	0.16	Tolerated	3.77	5	-1	0	5.3	-58.04
c.3380G>C	G1127A	Likely Benign		Conflicting		4	6-33443932-G-C	4	2.68e-6	-5.949	Likely Benign	0.080	Likely Benign	Likely Benign	0.164	Likely Benign										-0.43	Neutral	0.001	Benign	0.002	Benign	4.83	Benign	1.00	Tolerated	4.32	4	1	0	2.2	14.03
c.603T>G	D201E (3D Viewer)	Likely Benign	PH	Conflicting		2	6-33435245-T-G	20	1.24e-5	-2.640	Likely Benign	0.406	Ambiguous	Likely Benign	0.165	Likely Benign	0.42	Likely Benign	0.2	1.99	Ambiguous	1.21	Ambiguous	0.23	Likely Benign	-0.69	Neutral	0.633	Possibly Damaging	0.108	Benign	4.30	Benign	1.00	Tolerated	3.46	9	3	2	0.0	14.03	258.7	-24.8	0.9	0.1	-0.3	0.2						X		Uncertain	Asp201, an acidic residue located in the N-terminal loop before the first anti-parallel β sheet strand (res. Ile205-Pro208), is replaced by another acidic residue, glutamate. The carboxylate groups of both Asp201 and Glu201 side chains form hydrogen bonds with the hydroxyl group of Ser221 in the simulations. Due to its shorter side chain, Asp201 can also hydrogen bond with the backbone amide groups of neighboring loop residues Ser204 and Asp203. However, since the model ends abruptly at the N-terminus, no definite conclusions can be drawn from the simulations.
c.603T>A	D201E (3D Viewer)	Likely Benign	PH	Benign		1				-2.640	Likely Benign	0.406	Ambiguous	Likely Benign	0.165	Likely Benign	0.42	Likely Benign	0.2	1.99	Ambiguous	1.21	Ambiguous	0.23	Likely Benign	-0.69	Neutral	0.633	Possibly Damaging	0.108	Benign	4.30	Benign	1.00	Tolerated	3.46	9	3	2	0.0	14.03	258.7	-24.8	0.9	0.1	-0.3	0.2						X		Uncertain	Asp201, an acidic residue located in the N-terminal loop before the first anti-parallel β sheet strand (res. Ile205-Pro208), is replaced by another acidic residue, glutamate. The carboxylate groups of both Asp201 and Glu201 side chains form hydrogen bonds with the hydroxyl group of Ser221 in the simulations. Due to its shorter side chain, Asp201 can also hydrogen bond with the backbone amide groups of neighboring loop residues Ser204 and Asp203. However, since the model ends abruptly at the N-terminus, no definite conclusions can be drawn from the simulations.
c.3354C>A	S1118R	Likely Benign		Uncertain		1				-2.670	Likely Benign	0.553	Ambiguous	Likely Benign	0.166	Likely Benign										-0.74	Neutral	0.034	Benign	0.023	Benign	5.17	Benign	0.05	Affected	4.32	2	-1	0	-3.7	69.11
c.3494C>T	S1165L	Likely Benign		Conflicting		2				-2.984	Likely Benign	0.793	Likely Pathogenic	Ambiguous	0.166	Likely Benign										-2.01	Neutral	0.998	Probably Damaging	0.992	Probably Damaging	2.60	Benign	0.33	Tolerated	3.88	3	-3	-2	4.6	26.08																10.1016/j.ajhg.2020.11.011
c.3405G>C	K1135N	Likely Benign		Uncertain		1				-5.715	Likely Benign	0.960	Likely Pathogenic	Likely Pathogenic	0.166	Likely Benign										-0.97	Neutral	0.411	Benign	0.321	Benign	5.43	Benign	0.07	Tolerated	4.32	2	1	0	0.4	-14.07
c.3721C>A	L1241M		Coiled-coil	Uncertain		1				-5.881	Likely Benign	0.782	Likely Pathogenic	Likely Benign	0.167	Likely Benign										-1.43	Neutral	1.000	Probably Damaging	0.999	Probably Damaging	1.65	Pathogenic	0.00	Affected			4	2	-1.9	18.03
c.485G>A	R162H			Uncertain		1	6-33432782-G-A	2	1.24e-6	-9.730	Likely Pathogenic	0.480	Ambiguous	Likely Benign	0.167	Likely Benign										-1.13	Neutral	0.957	Probably Damaging	0.513	Possibly Damaging	4.03	Benign	0.12	Tolerated	3.74	4	2	0	1.3	-19.05
c.2854G>A	G952S	Likely Benign		Conflicting		2	6-33443406-G-A	2	1.24e-6	-6.190	Likely Benign	0.077	Likely Benign	Likely Benign	0.167	Likely Benign										0.19	Neutral	0.000	Benign	0.002	Benign	3.31	Benign	0.07	Tolerated	3.77	5	1	0	-0.4	30.03
c.2324G>C	R775P	Likely Benign		Benign		1				-5.072	Likely Benign	0.452	Ambiguous	Likely Benign	0.168	Likely Benign										-0.79	Neutral	0.971	Probably Damaging	0.944	Probably Damaging	4.13	Benign	0.07	Tolerated	3.64	6	-2	0	2.9	-59.07
c.3055C>T	R1019C	Likely Pathogenic		Conflicting		2	6-33443607-C-T	10	6.19e-6	-7.386	In-Between	0.646	Likely Pathogenic	Likely Benign	0.168	Likely Benign										-4.00	Deleterious	0.999	Probably Damaging	0.880	Possibly Damaging	2.36	Pathogenic	0.00	Affected	3.77	5	-4	-3	7.0	-53.05																10.1016/j.ajhg.2020.11.011
c.13C>G	R5G	Likely Benign		Uncertain		1				-3.639	Likely Benign	0.150	Likely Benign	Likely Benign	0.169	Likely Benign										-0.16	Neutral	0.013	Benign	0.003	Benign	4.12	Benign	0.00	Affected	4.32	1	-2	-3	4.1	-99.14
c.311G>T	R104L	Likely Benign		Benign		1	6-33432176-G-T	1	6.20e-7	-3.563	Likely Benign	0.578	Likely Pathogenic	Likely Benign	0.170	Likely Benign										-1.38	Neutral	0.001	Benign	0.002	Benign	4.05	Benign	0.00	Affected	4.32	1	-2	-3	8.3	-43.03
c.3820C>T	R1274C			Uncertain		1	6-33447868-C-T			-6.467	Likely Benign	0.439	Ambiguous	Likely Benign	0.170	Likely Benign										-5.22	Deleterious	1.000	Probably Damaging	0.996	Probably Damaging	2.46	Pathogenic	0.00	Affected	3.77	5	-4	-3	7.0	-53.05
c.2131C>G	L711V (3D Viewer)	Likely Pathogenic	GAP	Uncertain		1	6-33441596-C-G	1	6.20e-7	-10.045	Likely Pathogenic	0.709	Likely Pathogenic	Likely Benign	0.170	Likely Benign	3.48	Destabilizing	0.1	2.22	Destabilizing	2.85	Destabilizing	1.40	Destabilizing	-2.59	Deleterious	0.992	Probably Damaging	0.970	Probably Damaging	3.34	Benign	0.00	Affected	3.50	9	1	2	0.4	-14.03
c.3595G>A	E1199K		Coiled-coil	Uncertain		1	6-33446587-G-A	1	6.20e-7	-10.853	Likely Pathogenic	0.954	Likely Pathogenic	Ambiguous	0.171	Likely Benign										-2.26	Neutral	1.000	Probably Damaging	0.995	Probably Damaging	2.52	Benign	0.00	Affected	3.77	5	0	1	-0.4	-0.94
c.3197C>T	P1066L	Likely Benign		Likely Benign		1	6-33443749-C-T	14	8.71e-6	-5.478	Likely Benign	0.092	Likely Benign	Likely Benign	0.173	Likely Benign										-3.68	Deleterious	0.996	Probably Damaging	0.903	Possibly Damaging	2.72	Benign	0.00	Affected	4.32	2	-3	-3	5.4	16.04
c.3308G>T	R1103L	Likely Benign		Uncertain		1	6-33443860-G-T			-2.330	Likely Benign	0.205	Likely Benign	Likely Benign	0.173	Likely Benign										-2.35	Neutral	0.002	Benign	0.005	Benign	2.44	Pathogenic	0.02	Affected	3.77	5	-3	-2	8.3	-43.03
c.2245C>T	R749W			Likely Benign		1	6-33441710-C-T	3	1.86e-6	-7.647	In-Between	0.338	Likely Benign	Likely Benign	0.173	Likely Benign										-2.62	Deleterious	1.000	Probably Damaging	0.998	Probably Damaging	2.59	Benign	0.00	Affected	4.32	2	2	-3	3.6	30.03
c.3661C>T	R1221W	Likely Pathogenic	Coiled-coil	Conflicting		3	6-33446653-C-T	1	6.20e-7	-10.938	Likely Pathogenic	0.651	Likely Pathogenic	Likely Benign	0.174	Likely Benign										-4.57	Deleterious	1.000	Probably Damaging	0.987	Probably Damaging	2.50	Benign	0.01	Affected	3.77	5	2	-3	3.6	30.03
c.2443C>T	R815C	Likely Pathogenic	SH3-binding motif	Uncertain		1	6-33442995-C-T	5	3.10e-6	-9.373	Likely Pathogenic	0.828	Likely Pathogenic	Ambiguous	0.174	Likely Benign										-3.89	Deleterious	1.000	Probably Damaging	0.998	Probably Damaging	2.59	Benign	0.00	Affected	4.32	4	-4	-3	7.0	-53.05
c.2560C>T	R854C	Likely Benign		Uncertain		1	6-33443112-C-T	3	1.86e-6	-5.082	Likely Benign	0.170	Likely Benign	Likely Benign	0.174	Likely Benign										-2.48	Neutral	1.000	Probably Damaging	0.947	Probably Damaging	4.05	Benign	0.01	Affected	3.88	3	-3	-4	7.0	-53.05
c.2444G>T	R815L	Likely Pathogenic	SH3-binding motif	Uncertain		1				-8.546	Likely Pathogenic	0.865	Likely Pathogenic	Ambiguous	0.175	Likely Benign										-3.06	Deleterious	0.999	Probably Damaging	0.997	Probably Damaging	2.63	Benign	0.03	Affected	4.32	4	-2	-3	8.3	-43.03
c.3557C>T	S1186L		Coiled-coil	Uncertain		1	6-33444592-C-T			-4.829	Likely Benign	0.923	Likely Pathogenic	Ambiguous	0.177	Likely Benign										-2.58	Deleterious	0.998	Probably Damaging	0.992	Probably Damaging	2.65	Benign	0.04	Affected	3.82	4	-3	-2	4.6	26.08
c.1604G>C	S535T (3D Viewer)	Likely Benign	GAP	Benign		1	6-33438847-G-C	14	8.67e-6	-3.886	Likely Benign	0.069	Likely Benign	Likely Benign	0.177	Likely Benign	0.45	Likely Benign	0.1	-0.27	Likely Benign	0.09	Likely Benign	0.17	Likely Benign	-0.81	Neutral	0.000	Benign	0.001	Benign	-1.25	Pathogenic	0.25	Tolerated	3.37	35	1	1	0.1	14.03	201.3	-17.3	-0.1	0.7	-0.2	0.1						X		Potentially Benign	Ser535 is located near the terminal end of an α-helix (res. Ala533-Val560) close to the membrane interface. In the WT simulations, the hydroxyl side chain of Ser535 forms hydrogen bonds with nearby residues (e.g., His539, Glu538) without any specific interactions. These hydrogen bonds disrupt the structure of the terminal end of the α-helix (Ala533-Ser535), causing it to weaken or unfold during the WT simulations. In the variant simulations, Thr535, a hydrophilic residue with a hydroxyl group of almost the same size as Ser, interacts more frequently with the preceding loop residues (e.g., Thr532, Cys531) due to its longer side chain. Regardless, the residue swap is tolerated in the simulations with no negative effects. However, due to its location near the SynGAP-membrane interface, the effect of the residue swap cannot be fully addressed using the SynGAP solvent-only simulations.	10.1016/j.ajhg.2020.11.011
c.3835G>A	A1279T	Likely Benign		Uncertain		2	6-33447883-G-A	2	1.29e-6	-4.871	Likely Benign	0.071	Likely Benign	Likely Benign	0.178	Likely Benign										-0.30	Neutral	0.001	Benign	0.000	Benign	2.71	Benign	0.09	Tolerated	3.77	5	1	0	-2.5	30.03
c.1339G>C	V447L (3D Viewer)	Likely Benign	GAP	Uncertain		1				-5.136	Likely Benign	0.491	Ambiguous	Likely Benign	0.180	Likely Benign	-1.13	Ambiguous	0.1	0.54	Ambiguous	-0.30	Likely Benign	0.03	Likely Benign	-0.29	Neutral	0.947	Possibly Damaging	0.851	Possibly Damaging	3.61	Benign	0.90	Tolerated	3.37	32	1	2	-0.4	14.03
c.3397A>G	I1133V	Likely Benign		Benign		1	6-33443949-A-G	22	1.48e-5	-3.362	Likely Benign	0.067	Likely Benign	Likely Benign	0.180	Likely Benign										0.06	Neutral	0.007	Benign	0.007	Benign	5.47	Benign	0.58	Tolerated	4.32	3	4	3	-0.3	-14.03																10.1016/j.ajhg.2020.11.011
c.2681G>A	G894E	Likely Benign		Uncertain		1	6-33443233-G-A	6	3.72e-6	-5.377	Likely Benign	0.859	Likely Pathogenic	Ambiguous	0.180	Likely Benign										-2.07	Neutral	1.000	Probably Damaging	1.000	Probably Damaging	2.68	Benign	0.01	Affected	4.32	4	0	-2	-3.1	72.06
c.1428C>G	F476L (3D Viewer)		GAP	Uncertain		2	6-33438460-C-G	4	2.48e-6	-10.109	Likely Pathogenic	0.994	Likely Pathogenic	Likely Pathogenic	0.180	Likely Benign	1.00	Ambiguous	0.1	1.04	Ambiguous	1.02	Ambiguous	0.75	Ambiguous	-1.10	Neutral	0.997	Probably Damaging	0.978	Probably Damaging	3.53	Benign	0.60	Tolerated	3.40	22	2	0	1.0	-34.02	235.9	16.1	0.0	0.1	-0.2	0.0	X							Potentially Benign	In the WT simulations, the phenyl ring of Phe476, located at the end of an α-helix (res. Ala461-Phe476), packs with the hydrophobic side chains of Leu482 and Ile483. Additionally, Phe476 stacks with the Arg475 side chain on the preceding α-α loop connecting the two α-helices (res. Ala461-Phe476 and res. Leu489-Glu519) near the GAP-Ras interface.In the variant simulations, Leu476 can maintain hydrophobic packing with neighboring residues, although not as efficiently as the phenylalanine in the WT system. The absence of Phe476/Arg475 stacking weakens the integrity of the α-helix end in the variant simulations. Nonetheless, no large-scale adverse effects are observed in the simulations. Lastly, the potential effect of the residue swap on SynGAP-Ras complex formation or GTPase activation cannot be fully addressed using the SynGAP solvent-only simulations.
c.2218C>T	R740W			Uncertain		2	6-33441683-C-T	6	3.72e-6	-8.561	Likely Pathogenic	0.168	Likely Benign	Likely Benign	0.180	Likely Benign										-3.09	Deleterious	1.000	Probably Damaging	0.938	Probably Damaging	2.52	Benign	0.01	Affected	4.32	2	2	-3	3.6	30.03
c.455G>A	R152Q			Uncertain		1	6-33432752-G-A	5	3.14e-6	-10.336	Likely Pathogenic	0.989	Likely Pathogenic	Likely Pathogenic	0.181	Likely Benign										-2.34	Neutral	0.997	Probably Damaging	0.968	Probably Damaging	3.89	Benign	0.00	Affected	3.61	5	1	1	1.0	-28.06
c.2561G>A	R854H	Likely Benign		Uncertain		1	6-33443113-G-A	4	2.48e-6	-3.686	Likely Benign	0.094	Likely Benign	Likely Benign	0.183	Likely Benign										-1.38	Neutral	0.997	Probably Damaging	0.899	Possibly Damaging	4.07	Benign	0.04	Affected	3.88	3	2	0	1.3	-19.05
c.2115G>C	K705N (3D Viewer)	Likely Pathogenic	GAP	Likely Pathogenic		1				-9.767	Likely Pathogenic	0.925	Likely Pathogenic	Ambiguous	0.183	Likely Benign	0.74	Ambiguous	0.0	0.37	Likely Benign	0.56	Ambiguous	0.44	Likely Benign	-3.12	Deleterious	0.996	Probably Damaging	0.876	Possibly Damaging	3.37	Benign	0.02	Affected	3.47	10	1	0	0.4	-14.07	221.4	-20.2	0.0	0.0	0.0	0.1						X		Uncertain	The amino side chain of Lys705, located at the end and outer surface of an α-helix (res. Thr704-Gly712), does not form any interactions in the WT simulations. In the variant simulations, the carboxamide side chain of Asn705 briefly forms a salt bridge with Glu706. However, there is no apparent difference between the systems. Due to the model ending abruptly at the C-terminus, no definite conclusions can be drawn based on the simulations.
c.2522T>C	V841A			Uncertain		1	6-33443074-T-C	3	1.86e-6	-8.152	Likely Pathogenic	0.901	Likely Pathogenic	Ambiguous	0.183	Likely Benign										-2.13	Neutral	0.992	Probably Damaging	0.989	Probably Damaging	2.57	Benign	0.02	Affected	3.77	5	0	0	-2.4	-28.05
c.323A>G	K108R	Likely Benign		Uncertain		1	6-33432188-A-G	6	3.72e-6	-2.892	Likely Benign	0.148	Likely Benign	Likely Benign	0.184	Likely Benign										0.37	Neutral	0.993	Probably Damaging	0.956	Probably Damaging	4.22	Benign	1.00	Tolerated	3.61	5	3	2	-0.6	28.01
c.491G>A	R164Q			Uncertain		1	6-33432788-G-A	2	1.24e-6	-11.208	Likely Pathogenic	0.600	Likely Pathogenic	Likely Benign	0.184	Likely Benign										-1.86	Neutral	0.957	Probably Damaging	0.342	Benign	3.82	Benign	0.00	Affected	3.74	4	1	1	1.0	-28.06
c.3862A>G	K1288E			Uncertain		1	6-33447910-A-G	5	3.22e-6	-2.751	Likely Benign	0.407	Ambiguous	Likely Benign	0.185	Likely Benign										-3.27	Deleterious	0.979	Probably Damaging	0.973	Probably Damaging	2.13	Pathogenic	0.00	Affected	3.77	5	1	0	0.4	0.94
c.2852A>G	H951R	Likely Benign		Likely Pathogenic		1				-4.964	Likely Benign	0.125	Likely Benign	Likely Benign	0.185	Likely Benign										-1.08	Neutral	0.048	Benign	0.029	Benign	5.46	Benign	0.24	Tolerated	3.77	5	2	0	-1.3	19.05
c.2741A>T	D914V	Likely Benign		Uncertain		1				-4.260	Likely Benign	0.723	Likely Pathogenic	Likely Benign	0.187	Likely Benign										-2.24	Neutral	0.999	Probably Damaging	0.986	Probably Damaging	2.64	Benign	0.01	Affected	3.77	5	-3	-2	7.7	-15.96
c.3364G>A	G1122S	Likely Benign		Benign/Likely benign		2	6-33443916-G-A	27	1.79e-5	-4.880	Likely Benign	0.072	Likely Benign	Likely Benign	0.189	Likely Benign										-0.08	Neutral	0.022	Benign	0.006	Benign	4.89	Benign	0.92	Tolerated	3.77	5	1	0	-0.4	30.03
c.892C>T	P298S (3D Viewer)	Likely Benign	C2	Benign		1	6-33437797-C-T	5	3.10e-6	-6.342	Likely Benign	0.144	Likely Benign	Likely Benign	0.189	Likely Benign	1.38	Ambiguous	0.2	1.41	Ambiguous	1.40	Ambiguous	0.58	Ambiguous	-1.20	Neutral	0.991	Probably Damaging	0.898	Possibly Damaging	2.03	Pathogenic	0.85	Tolerated	3.39	20	-1	1	0.8	-10.04
c.76G>A	G26R	Likely Benign		Benign		1	6-33423485-G-A	3	1.86e-6	-2.946	Likely Benign	0.678	Likely Pathogenic	Likely Benign	0.189	Likely Benign										-2.22	Neutral	0.994	Probably Damaging	0.990	Probably Damaging	3.87	Benign	0.00	Affected	4.32	1	-3	-2	-4.1	99.14
c.484C>G	R162G	Likely Benign		Uncertain		1				-6.985	Likely Benign	0.664	Likely Pathogenic	Likely Benign	0.190	Likely Benign										-0.73	Neutral	0.487	Possibly Damaging	0.272	Benign	4.09	Benign	0.78	Tolerated	3.74	4	-2	-3	4.1	-99.14
c.407G>A	R136Q			Benign		1	6-33432704-G-A	13	9.17e-6	-11.146	Likely Pathogenic	0.950	Likely Pathogenic	Ambiguous	0.190	Likely Benign										-2.26	Neutral	0.957	Probably Damaging	0.342	Benign	3.52	Benign	0.01	Affected	3.61	5	1	1	1.0	-28.06
c.2206C>T	R736C			Conflicting		3	6-33441671-C-T	8	4.96e-6	-7.113	In-Between	0.120	Likely Benign	Likely Benign	0.190	Likely Benign										-2.06	Neutral	0.999	Probably Damaging	0.825	Possibly Damaging	2.48	Pathogenic	0.00	Affected	4.07	3	-4	-3	7.0	-53.05
c.2669G>C	R890P	Likely Benign		Likely Benign		2	6-33443221-G-C	28	1.74e-5	-1.931	Likely Benign	0.301	Likely Benign	Likely Benign	0.191	Likely Benign										-1.21	Neutral	0.999	Probably Damaging	0.977	Probably Damaging	4.02	Benign	0.28	Tolerated	4.32	4	0	-2	2.9	-59.07
c.2282G>A	R761Q	Likely Benign		Uncertain		1	6-33441747-G-A	11	6.81e-6	-4.187	Likely Benign	0.202	Likely Benign	Likely Benign	0.191	Likely Benign										-0.63	Neutral	0.996	Probably Damaging	0.878	Possibly Damaging	2.75	Benign	0.40	Tolerated	3.99	5	1	1	1.0	-28.06
c.1042G>A	V348M (3D Viewer)		C2	Uncertain		1				-7.076	In-Between	0.546	Ambiguous	Likely Benign	0.191	Likely Benign	-1.19	Ambiguous	0.1	0.72	Ambiguous	-0.24	Likely Benign	0.76	Ambiguous	-1.62	Neutral	0.966	Probably Damaging	0.564	Possibly Damaging	1.58	Pathogenic	0.03	Affected	3.37	25	2	1	-2.3	32.06	253.8	-47.4	-0.3	0.1	0.2	0.1						X		Potentially Benign	The iso-propyl side chain of Val348, located in an anti-parallel β sheet strand (res. Gly341-Pro349), packs against multiple hydrophobic C2 domain residues (e.g., Leu353, Leu323, Leu402). In the variant simulations, the thioether side chain of Met348 can form similar interactions as valine due to its comparable hydrophobic profile. In fact, the thioether group of methionine can even stack favorably with the phenol ring of Tyr363 in the anti-parallel β sheet strand (res. Ala399-Ile411). Overall, the residue swap does not appear to cause negative effects on the protein structure based on the simulations.
c.1436G>A	R479Q (3D Viewer)	Likely Benign	GAP	Uncertain		1	6-33438468-G-A	7	4.34e-6	-7.109	In-Between	0.259	Likely Benign	Likely Benign	0.191	Likely Benign	0.54	Ambiguous	0.1	0.57	Ambiguous	0.56	Ambiguous	0.49	Likely Benign	-1.16	Neutral	1.000	Probably Damaging	0.991	Probably Damaging	3.42	Benign	0.31	Tolerated	3.39	32	1	1	1.0	-28.06
c.2525C>A	S842Y	Likely Pathogenic		Likely Pathogenic		1				-16.124	Likely Pathogenic	0.995	Likely Pathogenic	Likely Pathogenic	0.191	Likely Benign										-4.28	Deleterious	0.944	Possibly Damaging	0.676	Possibly Damaging	1.97	Pathogenic	0.00	Affected	3.77	5	-3	-2	-0.5	76.10
c.3920C>A	P1307Q	Likely Benign		Uncertain		1	6-33451794-C-A			-4.227	Likely Benign	0.114	Likely Benign	Likely Benign	0.192	Likely Benign										-0.88	Neutral	0.988	Probably Damaging	0.765	Possibly Damaging	2.82	Benign	0.03	Affected	3.77	5	0	-1	-1.9	31.01
c.2714G>A	R905H	Likely Benign		Uncertain		1	6-33443266-G-A	8	4.96e-6	-4.182	Likely Benign	0.457	Ambiguous	Likely Benign	0.192	Likely Benign										-1.11	Neutral	1.000	Probably Damaging	0.991	Probably Damaging	2.59	Benign	0.09	Tolerated	3.77	5	2	0	1.3	-19.05
c.1300G>A	V434I (3D Viewer)	Likely Benign	GAP	Uncertain		1	6-33438205-G-A	1	6.19e-7	-6.999	Likely Benign	0.129	Likely Benign	Likely Benign	0.192	Likely Benign	-0.04	Likely Benign	0.0	0.22	Likely Benign	0.09	Likely Benign	0.31	Likely Benign	-0.82	Neutral	0.947	Possibly Damaging	0.851	Possibly Damaging	3.53	Benign	0.18	Tolerated	3.37	29	4	3	0.3	14.03	246.7	-27.7	0.0	0.0	0.1	0.0						X		Potentially Benign	The iso-propyl side chain of Val434, located at the end of an α helix (res. Met414-Glu436), packs against hydrophobic residues in an interhelix space (e.g., Met430, Ala707, Leu711). In the variant simulations, the sec-butyl group of Ile434 is able to form the same hydrophobic interactions. Accordingly, the residue swap does not negatively affect the protein structure based on the simulations.
c.2619C>G	S873R			Uncertain		1	6-33443171-C-G	1	6.20e-7	-5.856	Likely Benign	0.976	Likely Pathogenic	Likely Pathogenic	0.192	Likely Benign										-2.74	Deleterious	0.997	Probably Damaging	0.995	Probably Damaging	2.67	Benign	0.06	Tolerated	3.77	5	0	-1	-3.7	69.11
c.3116T>C	I1039T	Likely Benign		Uncertain		1	6-33443668-T-C	12	7.43e-6	-2.465	Likely Benign	0.645	Likely Pathogenic	Likely Benign	0.193	Likely Benign										0.45	Neutral	0.004	Benign	0.008	Benign	2.75	Benign	0.10	Tolerated	3.77	5	-1	0	-5.2	-12.05
c.2548G>A	G850R	Likely Benign		Uncertain		1				-5.082	Likely Benign	0.398	Ambiguous	Likely Benign	0.194	Likely Benign										-0.07	Neutral	0.010	Benign	0.010	Benign	4.30	Benign	0.01	Affected	3.77	5	-3	-2	-4.1	99.14
c.1976C>T	S659F (3D Viewer)	Likely Pathogenic	GAP	Uncertain		1				-10.925	Likely Pathogenic	0.662	Likely Pathogenic	Likely Benign	0.194	Likely Benign	-0.81	Ambiguous	0.1	-0.25	Likely Benign	-0.53	Ambiguous	0.32	Likely Benign	-4.59	Deleterious	0.806	Possibly Damaging	0.171	Benign	3.39	Benign	0.05	Affected	3.38	28	-3	-2	3.6	60.10	221.3	-61.2	0.0	0.0	0.6	0.4						X		Potentially Benign	In the WT simulations, the hydroxyl group of Ser659, located in a kink in the middle of the long α-helix (res. Ser641-Glu666), forms a hydrogen bond with the carboxylate group of Glu656. However, the phenol ring of the Phe659 side chain cannot form a similar hydrogen bond. Instead, it interacts with the hydrophobic isopropyl side chain of Val555 from the opposing α-helix (res. Ala533-Val560). This residue swap may therefore cause issues during protein folding.
c.2713C>T	R905C			Conflicting		2	6-33443265-C-T	15	9.31e-6	-5.578	Likely Benign	0.723	Likely Pathogenic	Likely Benign	0.194	Likely Benign										-3.14	Deleterious	1.000	Probably Damaging	0.980	Probably Damaging	2.57	Benign	0.01	Affected	3.77	5	-4	-3	7.0	-53.05
c.2485G>A	E829K	Likely Pathogenic		Pathogenic		1				-7.527	In-Between	0.807	Likely Pathogenic	Ambiguous	0.194	Likely Benign										-2.65	Deleterious	0.994	Probably Damaging	0.900	Possibly Damaging	2.27	Pathogenic	0.00	Affected	3.77	5	0	1	-0.4	-0.94
c.2255C>T	S752L	Likely Benign		Uncertain		2	6-33441720-C-T	6	3.72e-6	-3.386	Likely Benign	0.182	Likely Benign	Likely Benign	0.195	Likely Benign										-2.09	Neutral	0.993	Probably Damaging	0.641	Possibly Damaging	1.51	Pathogenic	0.01	Affected	3.99	5	-3	-2	4.6	26.08
c.2945A>G	Y982C	Likely Benign		Likely Benign		1	6-33443497-A-G	2	1.24e-6	-6.256	Likely Benign	0.746	Likely Pathogenic	Likely Benign	0.195	Likely Benign										-1.67	Neutral	0.997	Probably Damaging	0.923	Probably Damaging	3.87	Benign	0.00	Affected	4.32	1	0	-2	3.8	-60.04
c.1998G>C	E666D (3D Viewer)	Likely Pathogenic	GAP	Uncertain		1				-8.820	Likely Pathogenic	0.704	Likely Pathogenic	Likely Benign	0.197	Likely Benign	0.88	Ambiguous	0.0	0.37	Likely Benign	0.63	Ambiguous	1.05	Destabilizing	-2.69	Deleterious	0.992	Probably Damaging	0.603	Possibly Damaging	3.43	Benign	0.06	Tolerated	3.38	28	3	2	0.0	-14.03	237.2	16.5	0.0	0.0	-0.3	0.1		X						Potentially Pathogenic	The carboxylate group of Glu666, located on the α-helix (res. Ser641-Glu666), is involved in a highly coordinated hydrogen-bonding network between residues from two α-helices (res. Ser641-Glu666 and res. Arg563-Glu578) and from the α-α loop connecting the two α-helices (res. Ser641-Glu666 and res. Leu685-Val699), such as Lys566, Thr672, and Asn669, in the WT simulations. In the variant simulations, the shorter side chain of Asp666 cannot maintain these interactions as efficiently as Glu666 in the WT, resulting in a less coordinated hydrogen-bond network.
c.86T>C	M29T	Likely Benign		Uncertain		1				-2.167	Likely Benign	0.122	Likely Benign	Likely Benign	0.199	Likely Benign										-0.37	Neutral	0.018	Benign	0.184	Benign	4.33	Benign	0.00	Affected	4.32	1	-1	-1	-2.6	-30.09
c.2668C>T	R890C			Benign		1	6-33443220-C-T	9	5.58e-6	-5.786	Likely Benign	0.402	Ambiguous	Likely Benign	0.200	Likely Benign										-3.38	Deleterious	1.000	Probably Damaging	0.971	Probably Damaging	3.94	Benign	0.04	Affected	4.32	4	-4	-3	7.0	-53.05
c.2282G>C	R761P	Likely Benign		Uncertain		3	6-33441747-G-C	1	6.20e-7	-5.091	Likely Benign	0.640	Likely Pathogenic	Likely Benign	0.201	Likely Benign										-1.89	Neutral	0.999	Probably Damaging	0.968	Probably Damaging	2.69	Benign	0.38	Tolerated	3.99	5	0	-2	2.9	-59.07
c.1594A>C	T532P (3D Viewer)	Likely Benign	GAP	Benign		1				-2.143	Likely Benign	0.061	Likely Benign	Likely Benign	0.201	Likely Benign	-0.30	Likely Benign	0.2	0.06	Likely Benign	-0.12	Likely Benign	0.08	Likely Benign	-0.90	Neutral	0.005	Benign	0.008	Benign	-1.28	Pathogenic	0.18	Tolerated	3.37	35	0	-1	-0.9	-3.99	174.2	35.1	0.4	0.0	0.1	0.0						X		Potentially Benign	Thr532 is located on an α-α loop between the two α-helices (res. Gly502-Tyr518 and Ala533-Val560) facing the membrane. In the WT simulations, the hydroxyl group of Thr532 occasionally forms hydrogen bonds with the backbone atoms of other loop residues without any specific interaction. In the variant simulations, the Pro532 residue swap does not cause structural changes. Although hydrophilic residues seem more favorable in the loop, the pyrrolidine side chain of proline is well suited for unstructured protein regions such as loops. However, due to its location at the SynGAP-membrane interface, the effect of the residue swap cannot be fully addressed using the SynGAP solvent-only simulations.
c.3511G>A	A1171T	Likely Benign	Coiled-coil	Uncertain		1				-3.658	Likely Benign	0.149	Likely Benign	Likely Benign	0.201	Likely Benign										-0.48	Neutral	0.245	Benign	0.138	Benign	5.45	Benign	0.07	Tolerated	4.32	4	1	0	-2.5	30.03
c.3253C>T	R1085W			Uncertain		1	6-33443805-C-T	2	1.26e-6	-6.339	Likely Benign	0.821	Likely Pathogenic	Ambiguous	0.202	Likely Benign										-3.15	Deleterious	1.000	Probably Damaging	0.996	Probably Damaging	2.70	Benign	0.00	Affected	3.77	5	-3	2	3.6	30.03
c.1600T>C	S534P (3D Viewer)	Likely Benign	GAP	Uncertain		1	6-33438843-T-C	3	1.86e-6	-5.056	Likely Benign	0.265	Likely Benign	Likely Benign	0.203	Likely Benign	-0.40	Likely Benign	0.2	0.35	Likely Benign	-0.03	Likely Benign	0.47	Likely Benign	-3.81	Deleterious	0.993	Probably Damaging	0.993	Probably Damaging	3.32	Benign	0.05	Affected	3.37	35	-1	1	-0.8	10.04