Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

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.
J Mol Biol 1992 Aug 20
PMID:Crystal structure analysis, refinement and enzymatic reaction mechanism of N-carbamoylsarcosine amidohydrolase from Arthrobacter sp. at 2.0 A resolution. 138 45

Crystal structures of N-carbamoylsarcosine amidohydrolase (CSHase; EC 3.5.1.59) have been analyzed by X-ray diffraction methods with two different inhibitors bound to the active site at 2.28 A and 2.37 A resolution. The catalytic center of the enzyme could be identified on the basis of these structures. The four substrate binding sites are situated at the intersubunit interfaces of the compact dimers AB and CD of the homotetrameric enzyme. Both inhibitors inactivate the enzyme irreversibly through covalent binding of their aldehyde groups to the thiol group of the active-site cysteine residue Cys177. Within the identified substrate binding sites a number of residues from different subunits are involved in hydrogen bonding of the inhibitors. Two residues (Ala172 and Thr173) that form an unusual cis-peptide bond at the binding site are important components in fixing the examined inhibitors by hydrogen bonds. An electrochemical enzyme assay for CSHase was used to test the effect of inhibitors and substrate analogs on the enzyme's activity, revealing the high substrate specificity of CSHase. The intrinsic tryptophan fluorescence of CSHase increases strongly upon substrate and inhibitor binding. As most of the tryptophyl residues are located at the active sites, they are thus considerably affected by ligand binding. Fluorescence-detected stopped-flow measurements have been used to study the kinetics of glyoxylate and substrate binding to CSHase. Substrate and inhibitor binding could clearly be distinguished in the stopped-flow experiments. Inhibitor binding reveals at least three different elementary processes, whereas substrate binding is much faster and contains phases with different signs in amplitude.
J Mol Biol 1996 Oct 25
PMID:Crystallographic and fluorescence studies of ligand binding to N-carbamoylsarcosine amidohydrolase from Arthrobacter sp. 891 6

The N-carbamoyl-D-amino-acid amidohydrolase (D-NCAase) is used on an industrial scale for the production of D-amino acids. The crystal structure of D-NCAase was solved by multiple isomorphous replacement with anomalous scattering using xenon and gold derivatives, and refined to 1.95 A resolution, to an R-factor of 18.6 %. The crystal structure shows a four-layer alpha/beta fold with two six-stranded beta sheets packed on either side by two alpha helices. One exterior layer faces the solvent, whereas the other one is buried and involved in the tight intersubunit contacts. A long C-terminal fragment extends from a monomer to a site near a dyad axis, and associates with another monomer to form a small and hydrophobic cavity, where a xenon atom can bind. Site-directed mutagenesis of His129, His144 and His215 revealed strict geometric requirements of these conserved residues to maintain a stable conformation of a putative catalytic cleft. A region located within this cleft involving Cys172, Glu47, and Lys127 is proposed for D-NCAase catalysis and is similar to the Cys-Asp-Lys site of N-carbamoylsarcosine amidohydrolase. The homologous active-site framework of these enzymes with distinct structures suggests convergent evolution of a common catalytic mechanism.
J Mol Biol 2001 Feb 16
PMID:Crystal structure and site-directed mutagenesis studies of N-carbamoyl-D-amino-acid amidohydrolase from Agrobacterium radiobacter reveals a homotetramer and insight into a catalytic cleft. 1123 98