Crystal structure analysis, refinement and enzymatic reaction mechanism of N-carbamoylsarcosine amidohydrolase from Arthrobacter sp. at 2.0 A resolution.
N-carbamoylsarcosine amidohydrolase from Arthrobacter sp., a tetramer of polypeptides with 264 amino acid residues each, has been crystallized and its structure solved and refined at 2.0 A resolution, to a crystallographic R-factor of 18.6%. The crystals employed in the analysis contain one tetramer of 116,000 M(r) in the asymmetric unit. The structure determination proceeded by multiple isomorphous replacement, followed by solvent-flattening and density averaging about the local diads within the tetramer. In the final refined model, the root-mean-square deviation from ideality is 0.01 A for bond distances and 2.7 degrees for bond angles. The asymmetric unit consists of 7853 protein atoms, 431 water molecules and four sulfate ions bound into the putative active site clefts in each subunit. One subunit contains a central six-stranded parallel beta-pleated sheet packed by helices on both sides. On one side, two helices face the solvent, while two of the helices on the other side are buried in the tight intersubunit contacts. The catalytic center of the enzyme, tentatively identified by inhibitor binding, is located at the interface between two subunits and involves residues from both. It is suggested that the nucleophilic group involved in hydrolysis of the substrate is the thiol group of Cys117 and a nucleophilic addition-elimination mechanism is proposed.[1]References
- Crystal structure analysis, refinement and enzymatic reaction mechanism of N-carbamoylsarcosine amidohydrolase from Arthrobacter sp. at 2.0 A resolution. Romão, M.J., Turk, D., Gomis-Rüth, F.X., Huber, R., Schumacher, G., Möllering, H., Rüssmann, L. J. Mol. Biol. (1992) [Pubmed]
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