Sequence based mutation analysis of GBA

Subset of SNPs

The ten SNPs shown in the table below and highlighted in Figure 1 were chosen for the analysis in this task. It was tried to include SNPs all over the protein, in order to investigate the influence of mutations in several parts of the protein. The mutated residues forming hydrogenbonds with the active site of glucocerebrosidase are included, as these should result in either a mal- or nonfunctioning protein. It was not easy to find missense mutations only listed in dbSNP, as most of them were listed in HGMD, too.

Figure 1: Selected mutations highlighted on structure of 10GS. The different colors indicate, in which database the mutation is listed: dbSNP (blue), HGMD (red), both (green)

Nr.	SNP ID/Accession Number	Database	Position	Amino Acid Change	Codon Change	Remarks
1	CM081634	HGMD	49	Gly - Ser	cGGC-AGC
2	rs74953658, CM050263	dbSNP, HGMD	63	Asp-Asn	tGAC-AAC
3	rs1141820	dbSNP	99	His - Arg	CAC - CGC	suspected, status not validated
4	CM880035	HGMD	159	Arg - Gln	CGG-CAG	synonymos mutation at this position listed in dbSNP
5	rs80205046, CM041347	dbSNP, HGMD	221	Pro - Leu	CCC - CTC
6	rs74731340, CM970620	dbSNP, HGMD	310	Ser - Asn	AGT - AAT
7	CM880036	HGMD	409	Asn - Ser	AAC-AGC	most common mutation found in gaucher disease type 1 patients
8	CM960698	HGMD	426	Pro - Leu	CCC-CTC
9	rs80020805, CM052245	dbSNP, HGMD	455	Met - Val	cATG-GTG
10	rs113825752	dbSNP	509	Leu - Pro	CTT - CCT

[Note that the positions of the SNPs listed in HGMD in this table are different from the corresponding codon numbers, indicated in Task 5, as this position is relative to the sequence of GBA including the signal peptide (+ 39 residues).]

Physicochemical Properties and Changes

Figure 2: Comparison of the wildtype (red) and mutated (blue) amino acids mapped to the structure of 1OGS.

Mutation 1

The wildtype amino acid, Glycine is nonpolar, whereas the mutated amino acid Serin is polar. This different polarity, could be an indication, that the mutation is damaging. Looking at the structure, one can see, that this residue (pos. 10 in mature protein) is situated in a loop at the exterior of the protein. Therefore a substitution should not affect the function of the protein that much.

Mutation 2

...

Mutation 3

Mutation 4

Mutation 5

Mutation 6

Mutation 7

Mutation 8

Mutation 9

Mutation 10

Mutation Analysis

SIFT

Usage

• Webserver: http://sift.jcvi.org/www/SIFT_seq_submit2.html
• Input: sequence in fasta format, substitutions of interest (Example for mutation 1: G49S)

Results

Mutation Nr.	Prediction	Score	Sequence Conservation
1	tolerated	0.51	3.05
2	tolerated	0.06	3.05
3	tolerated	0.62	3.04
4	affect protein function	0.03	3.01
5	affect protein function	0.00	3.01
6	tolerated	0.54	3.01
7	affect protein function	0.05	3.02
8	tolerated	0.83	3.05
9	tolerated	0.12	3.01
10	affect protein function	0.01	3.09

Coming soon: Multiple alignment of sequence with its homologs + Conditional Probability Matrix

Polyphen2

Usage

• Webserver: http://genetics.bwh.harvard.edu/pph2/
• Input: sequence in fasta format, position and substitution of mutation

Results

The results of PolyPhen-2 (HumDiv) are listed in the table below. Three mutations (Numbers 3, 6 and 8) have been predicted to be harmless. Interestingly, the most common mutation found in gaucher disease patients is only classified as possibly damaging.

Mutation Nr.	Prediction	Score	Sensitivity	Specificity
1	probably damaging	0.997	0.40	0.98
2	probably damaging	1.000	0.00	1.00
3	benign	0.000	1.00	0.00
4	probably damaging	1.000	0.00	1.00
5	probably damaging	1.000	0.00	1.00
6	benign	0.100	0.94	0.85
7	possibly damaging	0.573	0.88	0.91
8	benign	0.001	0.99	0.15
9	probably damaging	0.999	0.14	0.99
10	probably damaging	0.978	0.75	0.96