期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：1980
卷号：77
期号：8
页码：4499-4503
DOI：10.1073/pnas.77.8.4499
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：Multiple mutants of Escherichia coli defective in penicillin-binding proteins (PBPs) were constructed, and into these strains Co1E1 plasmids carrying the genes for PBP-1a, -1b, or -3 were introduced. From these plasmid-carrying strains, PBP-1a and -1b were purified by ampicillin-Sepharose affinity chromatography and PBP-3 by cephalexin-Sepharose affinity chromatography. Improved purification was achieved by differential elution with NH2OH. Purified PBP-1b synthesized murein when added to the membrane fraction of a PBP-1b-defective mutant, which by itself failed to support murein synthesis in vitro. The PBP-1b preparation was able to synthesize murein from the lipid intermediate extracted with chloroform/methanol but was unable to utilize UDP-linked precursors for murein synthesis. Murein synthesis was inhibited by vancomysin, ristocetin, moenomycin, and enduracidin, but not by beta-lactam antibiotics. The synthesized murein was shown to contain crosslinked muropeptides. Their crosslinking was abolished by action of beta-lactam antibiotics. The PBP-1a and -3 preparations showed substantially no activity for murein synthesis in the same reaction system. None of the three PBPs showed D-alanine carboxypeptidase activity with UDP-N-acetylmuramoyl-pentapeptide as substrate or endopeptidase activity with bis(disaccharide-peptide) as substrate.
