OVEREXPRESSION, PURIFICATION, AND CHARACTERIZATION OF UDP-N-ACETYLMURAMYL - L-ALANINE LIGASE FROM ESCHERICHIA-COLI

UDP-N-acetylmuramyl: L-alanine ligase from Escherichia coli was overex pressed more than 600-fold and purified to near homogeneity. The purif ied enzyme was found to ligate L-alanine, L-serine, and glycine, as we ll as the nonnatural amino acid beta-chloro-L-alanine, to UDP-N-acetyl muramic acid. On the basis of (i) the specificity constants of the enz yme determined for L-alanine, L-serine, and glycine and (ii) the level s of these amino acids in the intracellular pool, it was calculated th at the rates of incorporation of L-serine and glycine into peptidoglyc an precursor metabolites could maximally amount to 0.1 and 0.5%, respe ctively, of the rate of L-alanine incorporation.