IDENTITY AND SOME PROPERTIES OF THE L-THREONINE ALDOLASE ACTIVITY MANIFESTED BY PURE 2-AMINO-3-KETOBUTYRATE LIGASE OF ESCHERICHIA-COLI

2-Amino-3-ketobutyrate ligase catalyzes the reversible, pyridoxal 5'-p hosphate-dependent condensation of glycine with acetyl CoA forming the unstable intermediate, 2-amino-3-ketobutyrate. Several independent li nes of evidence indicate that the pure protein obtained in the purific ation of this ligase from Escherichia coli also has L-threonine aldola se activity. The evidence includes: (a), a constant ratio of specific activities (aldolase/ligase) at all stages of purifying 2-amino-3-keto butyrate ligase to homogeneity; (b), the same rate of loss of aldolase and ligase activities during controlled heat inactivation of the pure protein at 60-degrees-C in the absence, as well as in the presence of acetyl CoA, a protective substrate; (c), ratios of the two enzymic ac tivities that are not significantly different during slow inactivation by iodoacetamide, with and without L-threonine added; (d), coincident rates of loss and essentially identical rates of recovery of aldolase activity and ligase activity during resolution of the holoenzyme with hydroxylamine followed by reconstitution with pyridoxal 5'-phosphate. No aldolase activity is observed with D-threonine as substrate and L- allothreonine is about 25% as effective as L-threonine. Whereas ligase activity has a sharp pH optimum at 7.5, the aldolase activity of this pure protein is maximal at pH 9.0. Comparative apparent K(m) values f or glycine (ligase) and L-threonine (aldolase) are 10 mM and 0.9 mM, r espectively, whereas corresponding respective V(max) values were found to be 2.5 mumol of CoA released/min per mg vs. 0.014 mumol of acetald ehyde formed (NADH oxidized)/min per mg.