Polymerization by transglycosylation in the biosynthesis of the peptidoglycan of Escherichia coli K 12 and its inhibition by antibiotics

Abstract

The biosynthesis of bacterial cell wall peptidoglycan is a complex process involving cytoplasmic and membrane steps [l] . N-Acetylglucosaminyl-N-acetylmuramyl-(pentapeptide)-pyrophosphoryl-undecaprenol is the last membrane precursor prior to polymerization which proceeds by transglycosylation (formation of the linear glycan strands) and transpeptidation (crosslinking of the peptide subunits) [l] . This membrane intermediate has been utilized in cellfree systems for the formation of peptidoglycan [2-l] , but the difficulty in conveniently isolating it in adequate amounts has greatly limited the direct investigation of the polymerizing enzymes. Most commonly, the in vitro synthesis of peptidoglycan is carried out by incubating the cytoplasmic precursors, UDP-iV-acetylmuramyl-pentapeptide and UDP-Nacetylglucosamine, with appropriate particulate preparations, crude cell walls or treated cells [3-lo] (see [l] for ref. before 1972). Under these conditions only the over-all course of the different membrane reactions is considered. However, a study of each membrane step in itself is essential for the understanding of the mode of action of certain antibiotics, for the development of the genetic analysis of these reactions and for the determination of the mechanisms involved in their regulation. Recently, the transpeptidation step, uncoupled from the other membrane reactions, has been successfully studied with artificial systems of donor and acceptor peptides [ 1 ,111. As far as we are aware, the transglycosylation step has not yet been directly investigated to any extent. The present paper describes a convenient in vitro