Probing the catalytic activity of a cell division-specific transpeptidase in vivo with beta-lactams.
Penicillin-binding protein 3 (PBP3; also called FtsI) is a transpeptidase that catalyzes cross-linking of the peptidoglycan cell wall in the division septum of Escherichia coli. To determine whether the catalytic activity of PBP3 is activated during division, we assayed acylation of PBP3 with three beta-lactams (cephalexin, aztreonam, and piperacillin) in growing cells. Acylation of PBP3 with cephalexin, but not aztreonam or piperacillin, appeared to be stimulated by cell division. Specifically, cephalexin acylated PBP3 about 50% faster in a population of dividing cells than in a population of filamentous cells in which division was inhibited by inactivation or depletion of FtsZ, FtsA, FtsQ, FtsW, or FtsN. However, in a simpler in vitro system using isolated membranes, acylation with cephalexin was not impaired by depletion of FtsW or FtsN. A conflicting previous report that the ftsA3(Ts) allele interferes with acylation of PBP3 was found to be due to the presence of a thermolabile PBP3 in the strain used in that study. The new findings presented here are discussed in light of the hypothesis that the catalytic activity of PBP3 is stimulated by interaction(s) with other division proteins. We suggest that there might be allosteric activation of substrate binding.
Eberhardt
Christian
C
Department of Microbiology, University of Iowa, Iowa City, Iowa 52242, USA.
Kuerschner
Lars
L
Weiss
David S
DS
D001426:Bacterial Proteins; D002352:Carrier Proteins; D002455:Cell Division; D002462:Cell Membrane / Q000378:metabolism; D004926:Escherichia coli / Q000166:cytology* / Q000201:enzymology* / Q000254:growth & development; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D006602:Hexosyltransferases / Q000235:genetics / Q000378:metabolism*; D009097:Multienzyme Complexes / Q000235:genetics / Q000378:metabolism*; D002267:Muramoylpentapeptide Carboxypeptidase; D046915:Penicillin-Binding Proteins; D046928:Peptidoglycan Glycosyltransferase; D010458:Peptidyl Transferases / Q000235:genetics / Q000378:metabolism*; D013696:Temperature; D047090:beta-Lactams / Q000378:metabolism* / Q000494:pharmacology
D001426:Bacterial Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; C478704:FtsI protein, E coli; D009097:Multienzyme Complexes; D046915:Penicillin-Binding Proteins; D047090:beta-Lactams; D010458:Peptidyl Transferases; D006602:Hexosyltransferases; D046928:Peptidoglycan Glycosyltransferase; D002267:Muramoylpentapeptide Carboxypeptidase