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J. Bacteriol. doi:10.1128/JB.00273-07
Copyright (c) 2007, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Unstable Escherichia coli L-forms revisited: growth requires peptidoglycan synthesis

Institut Jacques Monod (C.N.R.S., Université Paris 6, Université Paris 7), 75251 Paris Cedex 05, France; Centro de Biología Molecular "Severo Ochoa" CSIC-UAM, Campus de Cantoblanco, 28049 Madrid, Spain

^* To whom correspondence should be addressed. Email: dari{at}ijm.jussieu.fr.

	Abstract

Growing bacterial L-forms are reputed to lack peptidoglycan, although cell division is normally inseparable from septal peptidoglycan synthesis. To explore which cell division functions L-forms use, we established a protocol for quantitatively converting a culture of a wild type Escherichia coli K-12 strain overnight to a growing L-form-like state, using the -lactam cefsulodin, inhibitor of penicillin-binding proteins 1A and 1B. In rich hypertonic medium containing cefsulodin, all cells are spherical and osmosensitive, like classical L-forms. Surprisingly, however, mutant studies showed that colony formation requires d-glutamate, diaminopimelate and MurA activity, all of which are specific to peptidoglycan synthesis. HPLC analysis confirmed that these L-form-like cells contain peptidoglycan, with 7% of the normal amount. Moreover, the -lactam piperacillin, a specific inhibitor of the cell division protein PBP3, rapidly blocks cell division of these L-form-like cells. Similarly, penicillin-induced L-form-like cells, which grow only within the agar layer of rich hypertonic plates, also require d-glutamate, diaminopimelate and MurA activity. These results strongly suggest that cefsulodin- and penicillin-induced L-form-like cells of E. coli - and possibly all L-forms - have residual peptidoglycan synthesis which is essential for their growth, probably required for cell division.

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