Homology modelling Gaucher Disease

The object of this task was to apply homology modelling for predicting the tertiary structure of glycosylceramidase given its [sequence]. For this, we first selected different templates which were than used to derive the structure of glycosylceramidase using three different homology modeling tools, namely Modeller, SWISS-MODEL, and the I-TASSER server. The resulting models were evaluated using both quality assessment scores and the native crystal structure 1ogs. Technical details are reported in our protocol.

Template selection

We used HHsearch for searching the PDB for homologous templates. <xr id="tab:templates"/> lists some of the top-ranking templates. 2nt0_A is identical to the target 1ogs_A and was therefore excluded. Although all listed hits are homologous to the target (HHsearch probability > 97%), their sequence identity was below 30%. We therefore selected 2wnw_A (blue) as a close homolog, 2y24_A (green) as an intermediate homolog, and 3nco_A (yellow) as a more distant homolog. Note that the latter two templates to not cover the complete target which makes the homology modeling process harder. <figtable id="tab:templates">

Hit Nr	Template	Identity	Query HMM	Prob
> 80% sequence identity
1	2nt0_A	100.0	1-497	100.0
40% - 80% sequence identity

< 30% sequence identity
2	2wnw_A	28.0	36-496	100.0
3	3clw_A	14.0	64-495	100.0
4	2y24_A	18.0	66-495	100.0
5	3kl0_A	19.0	65-495	100.0
6	3zr5_A	17.0	65-494	100.0
7	3ik2_A	14.0	65-495	99.2
22	3nco_A	11.0	113-384	97.7
28	1egz_A	12.0	113-387	97.4

Homologs found by HHsearch. Bold: selected templated used for the following modelling. </figtable>

Modeller

Modeller is a popular tools for building models by the satisfaction of spatial restrains which are derived, for instance, from one or several target-template alignments. The alignments can come from any alignment or homology search tool, or they can be built by modeller itself. We created models for our target protein by (1) using a single template and employing Modeller to compute the alignment, (2) using a single template and the alignment from HHsearch, and (3) using multiple templates. The Model quality was assessed via the DOPE and DOPE z-score reported by Modeller as well as the QMEAN6 score from the SWISS-MODEL workspace. We compared the resulting models to the crystal structure 1ogs_A via the weighted all-atom RMSD score computed by SAP, as well as the TM-score, GDT_TS, and GTD_HA score which we computed by the program TMscore.

Single-template modeling

<xr id="tab:modeller-single-models"/> shows the resulting models. 2wnw_A produced the best looking model: all major secondary structure elements coincided with the native structure 1ogs_A (red). Only the target range 1-31 which was not covered by template (cf. table <xr id="tab:templates"/> resulted in some deviating loop regions (at the top right corner). Although 2y24_A is less conserved than 2wnw_A, the model came close to the native structure. 3nco_A shares the same TIM beta/alpha-barrel domain (the tube at the center) than 1ogs_A but is missing the Glycosyl hydrolase domain (the sheets at the right side) such that the model was less well structured in this region.

2wnw_A

2y24_A

3nco_A

Dope score per residue.

Models built by Modeller using single templates and alignments computed by Modeller itself. Red: 1ogs_A. </figtable>

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Template	DOPE	DOPE z-score	QMEAN6	RMSD	TM-score	GDT_TS	GTD_HA

2wnw_A	-55925	-0.471	0.689	1.006	0.824	0.661	0.479
2y24_A	-47194	0.777	0.376	1.222	0.550	0.294	0.223
3nco_A	-44033	1.224	0.139	2.158	0.252	0.093	0.043

</figtable>

Multiple-template modeling

2wnw_A-2y24_A-3kl0_A

2wnw_A-3nco_A

3ik2_A-3nco_A-1egz_A

Dope score per residue.

Template	DOPE	DOPE z-score	QMEAN6	RMSD	TM-score	GDT_TS	GTD_HA

2wnw_A-2y24_A-3kl0	-39084	1.930	0.314	1.987	0.465	0.277	0.179
2wnw_A-3nco_A	-46962	0.807	0.556	1.256	0.778	0.593	0.409
3ik2_A-3nco_A-1egz_A	-25391	0.3881	0.101	6.607	0.241	0.062	0.029