Properties of a novel D-stereospecific aminopeptidase from Ochrobactrum anthropi.
A novel aminopeptidase active toward D-amino acid-containing peptides, D-amino acid amides, and D-amino acid esters has been purified 2,800-fold to homogeneity from a bacterium Ochrobactrum anthropi SCRC C1-38, which had been isolated from soil. The enzyme has a molecular weight of about 122,000 and is composed of two identical subunits (Mr = 59,000). The optimal pH for activity was 8.0. It showed strict D-stereospecificity toward substrates including low molecular weight D-amino acid amides such as D-alanine amide, D-alpha-aminobutyric acid amides, and D-serine amide; D-alanine N-alkylamides such as D-alanine-p-nitroanilide, D-alanine benzylamide, and D-alanine n-butylamide; and peptides with a D-alanine at the NH2 terminus such as D-alanylglycine, D-alanylglycylglycine, D-alanyl-L-alanyl-L-alanine, and D-alanine oligomers. Generally, the enzyme did not act on substrates composed of L-amino acid at the NH2 terminus, although it showed low stereospecificity only toward substrates such as the methyl esters of L-alanine, L-serine, and L-alanine-p-nitroanilide. Comparing the Km and Vmax values for the major substrates, it is clear that the enzyme prefers peptides to amino acid arylamides or amino acid amides. The enzyme was tentatively named as "D-aminopeptidase." EDTA and divalent cations have no effect on the enzyme activity. The enzyme appears to be a thiol peptidase.
Sagami Chemical Research Center, Kanagawa, Japan.
Y
Y
Asano
A
A
Nakazawa
Y
Y
Kato
K
K
Kondo
D000421:Alcaligenes / Q000201:enzymology*; D000595:Amino Acid Sequence; D000626:Aminopeptidases / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007526:Isoelectric Point; D007700:Kinetics; D008670:Metals / Q000494:pharmacology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D013057:Spectrum Analysis; D013237:Stereoisomerism; D013696:Temperature
D008670:Metals; D000626:Aminopeptidases