Functional characterization of a novel ArgA from Mycobacterium tuberculosis.
The Mycobacterium tuberculosis gene Rv2747 encodes a novel 19-kDa ArgA that catalyzes the initial step in L-arginine biosynthesis, namely the conversion of L-glutamate to alpha-N-acetyl-L-glutamate. Initial velocity studies reveal that Rv2747 proceeds through a sequential kinetic mechanism, with K(m) values of 280 mM for L-glutamine and 150 microM for acetyl-coenzyme A and with a k(cat) value of 200 min(-1). Initial velocity studies with L-glutamate showed that even at concentrations of 600 mM, saturation was not observed. Therefore, only a k(cat)/K(m) value of 125 M(-1) min(-1) can be calculated. Inhibition studies reveal that the enzyme is strongly regulated by L-arginine, the end product of the pathway (50% inhibitory concentration, 26 microM). The enzyme was completely inhibited by 500 microM arginine, with a Hill coefficient of 0.60, indicating negatively cooperative binding of L-arginine.
Errey
James C
JC
Department of Biochemistry, Albert Einstein College of Medicine, Bronx, NY 10461, USA.
Blanchard
John S
JS
D000105:Acetyl Coenzyme A / Q000378:metabolism; D000123:Acetyltransferases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D000595:Amino Acid Sequence; D051046:Amino-Acid N-Acetyltransferase; D001120:Arginine / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005971:Glutamates / Q000378:metabolism; D018698:Glutamic Acid / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009169:Mycobacterium tuberculosis / Q000201:enzymology*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D004791:Enzyme Inhibitors; D005971:Glutamates; D018698:Glutamic Acid; D000105:Acetyl Coenzyme A; D001120:Arginine; D000123:Acetyltransferases; D051046:Amino-Acid N-Acetyltransferase; C016195:N-acetylglutamic acid