Role of Ser216 in the mechanism of action of membrane‐bound lytic transglycosylase B: Further evidence for substrate‐assisted catalysis

Lytic transglycosylases cleave the β‐(1 → 4)‐glycosidic bond in the bacterial cell wall heteropolymer peptidoglycan between the N ‐acetylmuramic acid (MurNAc) and N ‐acetylglucosamine (GlcNAc) residues with the concomitant formation of a 1,6‐anhydromuramoyl residue. Based on sequence alignments, Ser216 in Pseudomonas aeruginosa membrane‐bound lytic transglycosylase B (MltB) was targeted for replacement with alanine to delineate its role in the enzyme's mechanism of action. The specific activity of the Ser216 → Ala MltB derivative was less than 12% of that for the wild‐type enzyme, while its substrate binding affinity remained virtually unaltered. These data are in agreement with a role of Ser216 in orienting the N ‐acetyl group on MurNAc at the −1 subsite of MltB for its participation in a substrate‐assisted mechanism of action.