Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.5.1.4 (deaminase)
 5,113 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The gene encoding the D-stereospecific amino-acid amidase from Ochrobactrum anthropi SV3 was cloned and sequenced. Analysis of 7.3 kb of genomic DNA revealed the presence of six ORFs, one of which (daaA) encodes the D-amino-acid amidase. This enzyme, DaaA, is composed of 363 amino-acid residues (molecular mass 40 082 Da), and the deduced amino-acid sequence exhibits homology to alkaline D-peptidase from Bacillus cereus DF4-B (32% identity), DD-peptidase from Streptomyces R61 (29% identity), and other penicillin-recognizing proteins. The DaaA protein contains the typical SXXK, YXN, and H(K)XG active-site motifs identified in the penicillin-binding proteins and beta-lactamases. The daaA gene modified in the nucleotide sequence upstream from its start codon was overexpressed in Escherichia coli. The activity of the recombinant DaaA enzyme in cell-free extracts of E. coli was 33.6 U. mg-1 with D-phenylalaninamide as substrate, which is about 350-fold higher than in extracts of O. anthropi SV3. This enzyme was purified to electrophoretic homogeneity by ammonium sulfate fractionation and three column chromatography steps. On gel-filtration chromatography, DaaA appeared to be a monomer with a molecular mass of 40 kDa. It had maximal activity at 45 degrees C and pH 9.0, and was completely inactivated in the presence of phenylmethanesulfonyl fluoride or Zn2+. DaaA had hydrolyzing activity toward D-amino-acid amides with aromatic or hydrophobic side chains, but did not act on the substrates for the DD-peptidase and beta-lactamase, despite their sequence similarity to DaaA. The characteristics of the recombinant DaaA are similar to those found for the native enzyme partially purified from O. anthropi SV3.
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PMID:Gene cloning, nucleotide sequencing, and purification and characterization of the D-stereospecific amino-acid amidase from Ochrobactrum anthropi SV3. 1072 42