Difference between revisions of "Structure-based mutation analysis Gaucher Disease"
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File:scwrl_10.png|10: R44S |
File:scwrl_10.png|10: R44S |
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</gallery> |
</gallery> |
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| − | <caption> |
+ | <caption>Rotamers of SNPs from <xr id="tab:mutations"/>. Blue: wildtype; Red: rotamer SCWRL.</caption> |
</figure> |
</figure> |
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<figure id="fig:scwrl_ss"> |
<figure id="fig:scwrl_ss"> |
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| − | [[File:scwrl_ss.png|thumb| |
+ | [[File:scwrl_ss.png|thumb|150px|<caption>Seconary structure elements of 2nt0_A (grey) compared to secondary structure elements of models built by SCRWL.</caption>]] |
| + | </figure> |
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| + | |||
| + | == FoldX == |
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| + | We employed FoldX <ref name="scwrl">Qiang Wang, Adrian A. Canutescu, and Roland L. Dunbrack, Jr.(2008). [http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2682191/ SCWRL and MolIDE: Computer programs for side-chain conformation prediction and homology modeling]. Nat Protoc.</ref> for substituting the wildtype residues listed in <xr id="tab:mutations"/> by the corresponding mutatant residue which is chosen from a rotamer library. <xr id="fig:foldx"/> denotes the results. |
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| + | |||
| + | <figure id="fig:foldx"> |
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| + | <gallery perrow=5 widths="100"> |
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| + | File:foldx_1.png|1: H60R |
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| + | File:foldx_2.png|2: V172I |
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| + | File:foldx_3.png|3: E111K |
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| + | File:foldx_4.png|4: L197P |
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| + | File:foldx_5.png|5: W209R |
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| + | File:foldx_6.png|6: L470P |
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| + | File:foldx_7.png|7: W312C |
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| + | File:foldx_8.png|8: A384D |
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| + | File:foldx_9.png|9: D443N |
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| + | File:foldx_10.png|10: R44S |
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| + | </gallery> |
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| + | <caption>Rotamers of SNPs from <xr id="tab:mutations"/>. Blue: wildtype; Red: rotamer SCWRL; Orange: rotamer FoldX.</caption> |
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</figure> |
</figure> |
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Revision as of 21:23, 23 June 2012
Cystral structure
<figtable id="tab:mutations">
| PDB | Res [Å] | R value | Coverage | pH |
|---|---|---|---|---|
| 2nt0 | 1.80 | 0.18 | 96% (40-536) | 4.5 |
| 3gxi | 1.84 | 0.19 | 96% (40-536) | 5.5 |
| 2v3f | 1.95 | 0.15 | 96% (40-536) | 6.5 |
| 2v3d | 1.96 | 0.16 | 96% (40-536) | 6.5 |
| 1ogs | 2.00 | 0.18 | 96% (40-536) | 4.6 |
The 5 crystral structures of glycosylceramidase with the highest resolution. The physiological lysosomal pH value is 4.5. 2nt0 was selected for the analysis. </figtable>
Mutations
<figtable id="tab:mutations">
| Nr | Pos P04062 |
Pos 2nt0_A |
From | To | Disease causing |
|---|---|---|---|---|---|
| 1 | 99 | 60 | H | R | No |
| 2 | 211 | 172 | V | I | No |
| 3 | 150 | 111 | E | K | Yes |
| 4 | 236 | 197 | L | P | Yes |
| 5 | 248 | 209 | W | R | Yes |
| 6 | 509 | 470 | L | P | No |
| 7 | 351 | 312 | W | C | Yes |
| 8 | 423 | 384 | A | D | Yes |
| 9 | 482 | 443 | D | N | No |
| 10 | 83 | 44 | R | S | No |
Mutations used for the structure-based mutation analysis. </figtable>
<figure id="fig:mutations">
</figure>
SCWRL
We employed SCWRL <ref name="scwrl">Qiang Wang, Adrian A. Canutescu, and Roland L. Dunbrack, Jr.(2008). SCWRL and MolIDE: Computer programs for side-chain conformation prediction and homology modeling. Nat Protoc.</ref> for substituting the wildtype residues listed in <xr id="tab:mutations"/> by the corresponding mutatant residue which is chosen from a rotamer library. <xr id="fig:scwrl"/> denotes the results.
<figure id="fig:scwrl">
1: H60R
2: V172I
3: E111K
4: L197P
5: W209R
6: L470P
7: W312C
8: A384D
9: D443N
10: R44S
Rotamers of SNPs from <xr id="tab:mutations"/>. Blue: wildtype; Red: rotamer SCWRL. </figure>
None of rotamers chosen by SCWRL clashed with another side-chain or the backbone. We further could not observer changes in the secondary structure or the formation cavities. The hydrogen bonding network changed in case of mutation number 1, 5, 7, and 8 (cf. <xr id="tab:scwrl"/>). W209R introduces a hydrophilic arginine which forms a hydrogen bond to T180. Although not predicted by SCWRL, the arginine might impact the protein structure. W312C is located next to the active site (cf. <xr id="fig:mutations"/>) and there exists a hydrogen bond to E340. Substitution the hydrophobic tryptohphane by a hydrophlic cysteine in the vicinity of the active site might account for the disease-causing effect of this mutation.
<figtable id="tab:scwrl">
| Nr | Mutation | Wildtype | Mutatant | Clashes | Structural change | ||
|---|---|---|---|---|---|---|---|
| H-bonds | Hydrophobicity | H-bonds | Hydrophobicity | ||||
| 1 | H60R | T471 | Hydrophilic | G62 | Hydrophilic | No | No |
| 2 | V172I | Hydrophobic | Hydrophobic | No | No | ||
| 3 | E111K | Hydrophilic | Hydrophilic | No | No | ||
| 4 | L197P | Hydrophobic | Hydrophobic | No | No | ||
| 5 | W209R | Hydrophobic | T180 | Hydrophilic | No | No | |
| 6 | L470P | T482 | Hydrophobic | T482 | Hydrophobic | No | No |
| 7 | W312C | E340, C342, P316 | Hydrophobic | E340, C342 | Hydrophilic | No | No |
| 8 | A384D | Hydrophobic | V404 | Hydrophilic | No | No | |
| 9 | D443N | Hydrophilic | Hydrophilic | No | No | ||
| 10 | R44S | S13, Y487 | Hydrophilic | S13, Y487 | Hydrophilic | No | No |
Structure-based analysis of SNPs from <xr id="tab:mutations"/>. H-bonds: residues involved in forming hydrogen bonds (cut-off: 3.2 Å). </figtable>
We further noticed that SCRWL changed the backbone at some positions which led to different secondary structure assignments (<xr id="fig:scwrl_ss"/>). The positions at which the deviations could be observed were independent from the mutated sites.
<figure id="fig:scwrl_ss">
</figure>
FoldX
We employed FoldX <ref name="scwrl">Qiang Wang, Adrian A. Canutescu, and Roland L. Dunbrack, Jr.(2008). SCWRL and MolIDE: Computer programs for side-chain conformation prediction and homology modeling. Nat Protoc.</ref> for substituting the wildtype residues listed in <xr id="tab:mutations"/> by the corresponding mutatant residue which is chosen from a rotamer library. <xr id="fig:foldx"/> denotes the results.
<figure id="fig:foldx">
1: H60R
2: V172I
3: E111K
4: L197P
5: W209R
6: L470P
7: W312C
8: A384D
9: D443N
10: R44S
Rotamers of SNPs from <xr id="tab:mutations"/>. Blue: wildtype; Red: rotamer SCWRL; Orange: rotamer FoldX. </figure>
References
<references/>
