title: Structure of an aliphatic amidase from Geobacillus pallidus RAPc8.
abstract: The amidase from Geobacillus pallidus RAPc8, a moderate thermophile, is a member of the nitrilase superfamily and catalyzes the conversion of amides to the corresponding carboxylic acids and ammonia. It shows both amide-hydrolysis and acyl-transfer activities and also exhibits stereoselectivity for some enantiomeric substrates, thus making it a potentially important industrial catalyst. The crystal structure of G. pallidus RAPc8 amidase at a resolution of 1.9 A was solved by molecular replacement from a crystal belonging to the primitive cubic space group P4(2)32. G. pallidus RAPc8 amidase is homohexameric in solution and its monomers have the typical nitrilase-superfamily alpha-beta-beta-alpha fold. Association in the hexamer preserves the eight-layered alpha-beta-beta-alpha:alpha-beta-beta-alpha structure across an interface which is conserved in the known members of the superfamily. The extended carboxy-terminal tail contributes to this conserved interface by interlocking the monomers. Analysis of the small active site of the G. pallidus RAPc8 amidase suggests that access of a water molecule to the catalytic triad (Cys, Glu, Lys) side chains would be impeded by the formation of the acyl intermediate. It is proposed that another active-site residue, Glu142, the position of which is conserved in the homologues, acts as a general base to catalyse the hydrolysis of this intermediate. The small size of the substrate-binding pocket also explains the specificity of this enzyme for short aliphatic amides and its asymmetry explains its enantioselectivity.
authors: Department of Molecular and Cell Biology, University of Cape Town, Rondebosch, South Africa.
authors: Serah W
authors: SW
authors: Kimani
authors: Vinod B
authors: VB
authors: Agarkar
authors: Donald A
authors: DA
authors: Cowan
authors: Muhammed F R
authors: MF
authors: Sayed
authors: B Trevor
authors: BT
authors: Sewell
mesh_terms: D000581:Amidohydrolases / Q000737:chemistry*; D001407:Bacillus / Q000201:enzymology*; D001665:Binding Sites; D002384:Catalysis; D018360:Crystallography, X-Ray / Q000379:methods; D006868:Hydrolysis; D008956:Models, Chemical; D008958:Models, Molecular; D008968:Molecular Conformation; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D015203:Reproducibility of Results; D013237:Stereoisomerism; D013379:Substrate Specificity
chemical_list: D000581:Amidohydrolases; C019665:amidase