Homology based structure predictions BCKDHA

1.Calculation of models

To find similar structures to BCKDHA we ran HHsearch:
hhsearch -i query -d database -o output

It found the following 10 hits in the pdb70 database.

No	Hit	Prob	E-value	P-value	Score	SS	Cols	Query HMM	Template HMM	Identity
1	2bfd_A 2-oxoisovalerate dehydr	1.0	1	1	791.3	0.0	400	1-400	1-400 (400)	99%
2	1qs0_A 2-oxoisovalerate dehydr	1.0	1	1	571.5	0.0	349	32-382	52-407 (407)	39%
3	1w85_A Pyruvate dehydrogenase	1.0	1	1	530.8	0.0	356	8-382	6-362 (368)	34%
4	1umd_A E1-alpha, 2-OXO acid de	1.0	1	1	521.8	0.0	351	34-386	16-367 (367)	37%
5	2ozl_A PDHE1-A type I, pyruvat	1.0	1	1	482.7	0.0	331	46-380	25-356 (365)	27%
6	3l84_A Transketolase; TKT, str	1.0	1	1	85.4	0.0	133	161-297	113-252 (632)	21%
7	2r8o_A Transketolase 1, TK 1;	1.0	1	1	74.5	0.0	121	161-285	113-245 (669)	33%
8	2o1x_A 1-deoxy-D-xylulose-5-ph	1.0	1	1	74.2	0.0	127	161-287	122-254 (629)	18%
9	1gpu_A Transketolase; transfer	1.0	1	1	74.2	0.0	140	161-302	115-265 (680)	22%
10	3m49_A Transketolase; alpha-be	1.0	1	1	68.8	0.0	121	161-285	139-271 (690)	31%

> 60% sequence identity:
-2bfd_A
> 40% sequence identity:
< 40% sequence identity (ideally go towards 20%) :
-1qs0_A, 1umd_A, 1w85_A, 2r8o_A, 3m49_A, 2ozl_A, 1gpu_A, 3l84_A, 2o1x_A, -1w85_A

HHSearch has only hits with an identity higher than 60% or lower than 40%.

These are the templates we will work with:
> 60% sequence identity:
-2bfd_A
< 40% sequence identity (ideally go towards 20%) :
-2r8o_A

Modeller

MODELLER is used for homology or comparative modeling of protein three-dimensional structures.It calculates a model containing all non-hydrogen atoms. There are also many other tasks provided by MODELLER like de novo modeling of loops in protein structures, optimization of various models of protein structure with respect to a flexibly defined objective function, multiple alignment of protein sequences and/or structures, clustering, searching of sequence databases, comparison of protein structures, etc.[1]

A tutorial is provided on [2] and on [3]

To run modeller with more than one target we use the targets:
-1DTW:A 95%
-2BFE:A 94%
-2BFB:A 99%
-2bfd:A 99%
-3m49:A 31%
-1gpu:A 22%
-2o1x:A 18%

Protocol Modeller

SWISS-MODEL

SWISS-MODEL server page

To find protein structure homology models SWISS-MODEL can be used. As input it needs a protein sequence or a UniProt AC Code. Optional the template PDB-Id and the chain or a template file can be assigned. SWISS-MODEL is a fully automated protein structure homology-modeling server. It is accessible via the ExPASy web server, or from the program DeepView (Swiss Pdb-Viewer).
SWISS-MODEL

SWISS-MODEL server:

ID	link
2bfd_A	2bfd_A
2r8o_A	2r8o_A

iTasser

1w85_A

1w85_A

Prediction for 1w85_A

Sequ GVKTFQFPFAEQLEKVAEQFPTFQILNEEGEVVNEEAMPELSDEQLKELMRRMVYTRILDQRSISLNRQGRLGFYAPTAGQEASQIASHFALEKEDFILP
Pred cccccccccHHHcccccccccSSSSSccccccccccccccccHHHHHHHHHHHHHHHHHHHHHHHHHHccccccccccccHHHHHHHHHHHcccccSSSc
Conf 9988678803312122677876989899998888756799999999999999999999999999999678854562888879999999986799898980

Sequ GYRDVPQIIWHGLPLYQAFLFSRGHFHGNQIPEGVNVLPPQIIIGAQYIQAAGVALGLKMRGKKAVAITYTGDGGTSQGDFYEGINFAGAFKAPAIFVVQ
Pred ccHHHHHHHHccccHHHHHHHHcccccccccccccccccccccHHccHHHHHHHHHHHHHcccccSSSSSScccccccHHHHHHHHHHHHHcccSSSSSS
Conf 6226899998699899999973687767878999865377730415555899999999964989889999347621101599999999996799899982

Sequ NNRFAISTPVEKQTVAKTLAQKAVAAGIPGIQVDGMDPLAVYAAVKAARERAINGEGPTLIETLCFRYGPHTMSGDDPTRYRSKELENEWAKKDPLVRFR
Pred cccSSccccHHHHHccccHHHHHHcccccSSSSccccHHHHHHHHHHHHHHHHcccccSSSSSSSSSScccccccccccccccHHHHHHHHHcccHHHHH
Conf 7972403229877479878986221799858987957999999999999999828998899999976158657889975678999999988379099999

Sequ KFLEAKGLWSEEEENNVIEQAKEEIKEAIKKADETPKQKVTDLISIMFEELPFNLKEQYEIYKEKESK
Pred HHHHHcccccHHHHHHHHHHHHHHHHHHHHHHHHcccccHHHHHHHHcccccHHHHHHHHHHHHHHcc
Conf 99998799999999999999999999999999858998999998451038998799999999998549

This prediction is based on several templates fount by iTasser itself.

Protocol Swissmodel

2.Evaluation of models

As we want to compare the predicted models to the existing PDB entry we need the PDB file for 1U5B. Here only chain A is important, as the sequence for chain A was used to create the models. Therefore chain B has to be removed from the PDB file. Herefore we used a program names ExtractChains.pl provided by <ref>http://www.rosettacommons.org/guide/PDB+Manipulation+Scripts</ref>.

Swissmodel

Numeric evaluation

QMEAN4 global scores

QMEANscore4

2bfd_A	2r8o_A
0.67	0.271

QMEAN Z-Score

2bfd_A	2r8o_A
-1.604	-6.943
Z-Score plot1 2bfd_A	Z-Score plot1 2r8o_A
Z-Score plot2 2bfd_A	Z-Score plot2 2r8o_A

Score components

2bfd_A	2r8o_A
score components 2bfd_A	score components 2r8o_A

Local scores

2bfd_A	2r8o_A
Coloring by residue error 2bfd_A	Coloring by residue error 2r8o_A
Residue error plot 2bfd_A	Residue error plot 2r8o_A

Global scores: QMEAN4:

	2bfd_A		2r8o_A
Scoring function term	Raw score	Z-score	Raw score	Z-score
C_beta interaction energy	-162.66	0.54	-47.91	-1.49
All-atom pairwise energy	-10811.93	0.35	-2558.65	-1.98
Solvation energy	-27.04	-1.02	10.53	-4.08
Torsion angle energy	-75.78	-1.45	18.95	-4.99
QMEAN4 score	0.670	-1.60	0.271	-6.94

Local Model Quality Estimation

2bfd_A	2r8o_A
Local Model Quality Estimation 2bfd_A	Local Model Quality Estimation 2r8o_A