title: Structure determination through homology modelling and torsion-angle simulated annealing: application to a polysaccharide deacetylase from Bacillus cereus.
abstract: The structure of BC0361, a polysaccharide deacetylase from Bacillus cereus, has been determined using an unconventional molecular-replacement procedure. Tens of putative models of the C-terminal domain of the protein were constructed using a multitude of homology-modelling algorithms, and these were tested for the presence of signal in molecular-replacement calculations. Of these, only the model calculated by the SAM-T08 server gave a consistent and convincing solution, but the resulting model was too inaccurate to allow phase determination to proceed to completion. The application of slow-cooling torsion-angle simulated annealing (started from a very high temperature) drastically improved this initial model to the point of allowing phasing through cycles of model building and refinement to be initiated. The structure of the protein is presented with emphasis on the presence of a C(α)-modified proline at its active site, which was modelled as an α-hydroxy-L-proline.
authors: Department of Molecular Biology and Genetics, Democritus University of Thrace, University Campus, 68100 Alexandroupolis, Greece.
authors: Vasiliki E
authors: VE
authors: Fadouloglou
authors: Maria
authors: M
authors: Kapanidou
authors: Athanasia
authors: A
authors: Agiomirgianaki
authors: Sofia
authors: S
authors: Arnaouteli
authors: Vassilis
authors: V
authors: Bouriotis
authors: Nicholas M
authors: NM
authors: Glykos
authors: Michael
authors: M
authors: Kokkinidis
mesh_terms: D000465:Algorithms; D000581:Amidohydrolases / Q000737:chemistry*; D001409:Bacillus cereus / Q000201:enzymology*; D001426:Bacterial Proteins / Q000737:chemistry*; D020134:Catalytic Domain; D003460:Crystallization; D018360:Crystallography, X-Ray / Q000379:methods*; D056004:Molecular Dynamics Simulation / Q000592:standards*; D011392:Proline / Q000737:chemistry; D017433:Protein Structure, Secondary; D040681:Structural Homology, Protein
chemical_list: D001426:Bacterial Proteins; D011392:Proline; D000581:Amidohydrolases; C022427:polysaccharide deacetylase
keywords: molecular replacement; polysaccharide deacetylases; residue modifications; simulated annealing