Gene identification and characterization of the pyridoxine degradative enzyme alpha-(N-acetylaminomethylene)succinic acid amidohydrolase from Mesorhizobium loti MAFF303099.
We have found for the first time that a chromosomal gene, mlr6787, in Mesorhizobium loti encodes the pyridoxine degradative enzyme alpha-(N-acetylaminomethylene)succinic acid (AAMS) amidohydrolase. The recombinant enzyme expressed in Escherichia coli cells was homogeneously purified and characterized. The enzyme consisted of two subunits each with a molecular mass of 34,000+/-1,000 Da, and exhibited Km and kcat values of 53.7+/-6 microM and 307.3+/-12 min(-1), respectively. The enzyme required no cofactor or metal ion. The primary structure of AAMS amidohydrolase was elucidated for the first time here. The primary structure of the enzyme protein showed no significant identity to those of known hydrolase proteins and low homology to those of fluoroacetate dehalogenase (PDB code, 1Y37), haloalkane dehalogenase (1K5P), and aryl esterase (1VA4).
Yuan
Baiqiang
B
Department of Bioresources Science, Faculty of Agriculture, Kochi University, Nankoku, Kochi, Japan.
Yokochi
Nana
N
Yoshikane
Yu
Y
Ohnishi
Kouhei
K
Ge
Fei
F
Yagi
Toshiharu
T
D020561:Alphaproteobacteria / Q000201:enzymology*; D000581:Amidohydrolases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D015870:Gene Expression / Q000235:genetics*; D005798:Genes, Bacterial / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D006867:Hydrolases / Q000235:genetics*; D011736:Pyridoxine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D050644:Succinate-Semialdehyde Dehydrogenase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D013696:Temperature; D005680:gamma-Aminobutyric Acid / Q000031:analogs & derivatives / Q000737:chemistry / Q000235:genetics
D001426:Bacterial Proteins; D005680:gamma-Aminobutyric Acid; D050644:Succinate-Semialdehyde Dehydrogenase; C530614:2-(acetamidomethylene)succinate hydrolase, Mesorhizobium loti; D006867:Hydrolases; D000581:Amidohydrolases; D011736:Pyridoxine; C009338:succinic semialdehyde