[en] The low susceptibility of enterococci to beta-lactams is due to the activity of the low-affinity penicillin-binding protein 5 (PBP5). One important feature of PBP5 is its ability to substitute for most, if not all, penicillin-binding proteins when they are inhibited. That substitution activity was analyzed in Enterococcus hirae SL2, a mutant whose pbp5 gene was interrupted by the nisRK genes and whose PBP3 synthesis was submitted to nisin induction. Noninduced SL2 cells were unable to divide except when plasmid-borne pbp5 genes were present, provided that the PBP5 active site was functional. Potential protein-protein interaction sites of the PBP5 N-terminal module were mutagenized by site-directed mutagenesis. The T-167-L-184 region (designated site D) appeared to be an essential intramolecular site needed for the stability of the protein. Mutations made in the two globular domains present in the N-terminal module indicated that they were needed for the suppletive activity. The P-197-N-209 segment (site E) in one of these domains seemed to be particularly important, as single and double mutations reduced or almost completely abolished, respectively, the action of PBP5.
Alaedini, A., and R. A. Day. 1999. Identification of two penicillin-binding multienzyme complexes in Haemophilus influenzae. Biochem. Biophys. Res. Commun. 264:191-195.
Ausubel, F. M., R. Brant, R. E. Kingston, D. D. Moore, J. G. Seidman, J. A. Smith, and K. Struhl. 2001. Current protocols in molecular biology. John Wiley & Sons, New York, N.Y.
Bayles, K. W., E. W. Brunskill, J. J. Iandolo, L. L. Hruska, S. Huang, P. A. Pattee, B. K. Smiley, and R. E. Yasbin. 1994. A genetic and molecular characterization of the recA gene from Staphylococcus aureus. Gene 147:13-20.
Begg, K. J., and W. D. Donachie. 1985. Cell shape and division in Escherichia coli: experiments with shape and division mutants. J. Bacteriol. 163:615-622.
Biswas, I., A. Gruss, S. D. Ehrlich, and E. Maguin. 1993. High-efficiency gene inactivation and replacement system for gram-positive bacteria. J. Bacteriol. 175:3628-3635.
Buddelmeijer, N., and J. Beckwith. 2002. Assembly of cell division proteins at the E. coli cell center. Curr. Opin. Microbiol. 5:553-557.
Canepari, P., M. M. Lleo, R. Fontana, and G. Satta. 1986. In Streptococcus faecium penicillin-binding protein 5 alone is sufficient for growth at submaximal but not at maximal rate. J. Gen. Microbiol. 132:625-631.
Canepari, P., M. M. Lleo, R. Fontana, and G. Satta. 1987. Streptococcus faecium mutants that are temperature sensitive for cell growth and show alterations in penicillin-binding proteins. J. Bacteriol. 169;2432-2439.
Coyette, J., J. M. Ghuysen, and R. Fontana. 1980. The penicillin-binding proteins in Streptococcus faecalis ATCC 9790. Eur. J. Biochem. 110:445-456.
Coyette, J., A. Somzé, J. J. Biquet, and J. M. Ghuysen. 1983. Function of penicillin-binding protein 3 in Streptococcus faecium, p. 523-529. In R. Hakenbeck, J. V. Höltje, and H. Labischinski (ed.), The target of penicillin. Walter de Gruyter and Co., Berlin, Germany.
de Pedro, M. A., K. D. Young, J. V. Höltje, and H. Schwarz. 2003. Branching of Escherichia coli cells arises from multiple sites of inert peptidoglycan. J. Bacteriol. 185:1147-1152.
de Pedro, M. A., C. G. Grünfelder, and H. Schwarz. 2004. Restricted mobility of cell surface proteins in the polar regions of Escherichia coli. J. Bacteriol. 186:2594-2602.
de Ruyter, P. G., O. P. Kuipers, M. M. Beerthuyzen, I. van Alan-Boerrigter, and W. M. de Vos. 1996. Functional analysis of promoters in the nisin gene cluster of Lactococcus lactis. J. Bacteriol. 178:3434-3439.
de Ruyter, P. G., O. P. Kuipers, and W. M. de Vos. 1996. Controlled gene expression systems in Lactococcus lactis with the food-grade inducer nisin. Appl. Environ. Microbiol. 62:3662-3667.
de Vos, W. M. 1999. Gene expression systems for lactic acid bacteria. Curr. Opin. Microbiol. 2:289-295.
Di Lallo, G., M. Fagioli, D. Barionovi, P. Ghelardini, and L. Paolozzi. 2003. Use of a two-hybrid assay to study the assembly of a complex multicomponent protein machinery: bacterial septosome differentiation. Microbiology 149:3353-3359.
Duez, C., I. Thamm, F. Sapunaric, J. Coyette, and J. M. Ghuysen. 1998. The division and cell wall cluster of Enterococcus hirae S185. DNA Seq. 9:149-161.
Eisenberg, D., R. M. Weiss, and T. C. Terwilliger. 1982. The helical hydrophobic moment: a measure of amphiphilicity of a helix. Nature 299:371-374.
Eisenberg, D., E. Schwarz, M. Komaromy, and R. Wall. 1984. Analysis of membrane and surface protein sequences with the hydrophobic moment plot. J. Mol. Biol. 179:125-142.
El Kharroubi, A., P. Jacques, G. Piras, J. Van Beeumen, J. Coyette, and J. M. Ghuysen. 1991. The Enterococcus hirae R40 penicillin-binding protein 5 and the methicillin-resistant Staphylococcus aureus penicillin-binding protein 2′ are similar. Biochem. J. 280:463-469.
Errington, J., R. A. Daniel, and D. J. Scheffers. 2003. Cytokinesis in bacteria. Microbiol. Mol. Biol. Rev. 67:52-65.
Fontana, R., R. Cerini, P. Longoni, A. Grossato, and P. Canepari. 1983. Identification of a streptococcal penicillin-binding protein that reacts very slowly with penicillin. J. Bacteriol. 155:1343-1350.
Fontana, R., A. Grossato, L. Rossi, Y. R. Chang, and G. Satta. 1985. Transition from resistance to hypersusceptibility to β-lactam antibiotics associated with loss of a low-affinity penicillin-binding protein in a Streptococcus faecium mutant highly resistant to penicillin. Antimicrob. Agents Chemother. 28:678-683.
Gallet, X., B. Charloteaux, A. Thomas, and R. Brasseur. 2000. A fast method to predict protein interaction sites from sequences. J. Mol. Biol. 302:917-926.
Giesbrecht, P., J. Wecke, and B. Reinicke. 1976. On the morphogenesis of the cell wall of staphylococci. Int. Rev. Cytol. 44:225-318.
Goffin, C., and J. M. Ghuysen. 1998. Multimodular penicillin-binding proteins: an enigmatic family of orthologs and paralogs. Microbiol. Mol. Biol. Rev. 62:1079-1093.
Guerout-Fleury, A. M., K. Shazand, N. Frandsen, and P. Stragier. 1995. Antibiotic-resistance cassettes for Bacillus subtilis. Gene 167:335-336.
Higgins, M. L., and G. D. Shockman. 1970. Model of cell wall growth of Streptococcus faecalis. J. Bacteriol. 101:643-648.
Higgins, M. L., M. Ferrero, and L. Daneo-Moore. 1986. Relationship of shape to initiation of new sites of envelope growth in Streptococcus faecium cells treated with β-lactam antibiotics. J. Bacteriol. 167:562-569.
Höltje, J. 1996. A hypothetical holoenzyme involved in the replication of the murein sacculus of Escherichia coli. Microbiology 142:1911-1918.
Karimova, G., N. Dautin, and D. Ladant. 2005. Interaction network among Escherichia coli membrane proteins involved in cell division as revealed by bacterial two-hybrid analysis. J. Bacteriol. 187:2233-2243.
Kleerebezem, M., M. M. Beerthuyzen, E. E. Vaughan, W. M. de Vos, and O. P. Kuipers. 1997. Controlled gene expression systems for lactic acid bacteria: transferable nisin-inducible expression cassettes for Lactococcus, Leuconostoc, and Lactobacillus spp. Appl. Environ. Microbiol. 63:4581-4584.
Kruse, T., J. Bork-Jensen, and K. Gerdes. 2005. The morphogenetic MreBCD proteins of Escherichia coli form an essential membrane-bound complex. Mol. Microbiol. 55:79-89.
Kuipers, O. P., M. M. Beerthuyzen, P. G. de Ruyter, E. Luesink, and W. M. de Vos. 1995. Autoregulation of nisin biosynthesis in Lactococcus lactis by signal transduction. J. Biol. Chem. 270:27299-27304.
Lleo, M. M., P. Canepari, and G. Satta. 1990. Bacterial cell shape regulation: testing of additional predictions unique to the two-competing-sites models for peptidoglycan assembly and isolation of conditional rod-shaped mutants from some wild-type cocci. J. Bacteriol. 172:3758-3771.
Loureiro Dos Santos, A. L., and A. Chopin. 1987. Shotgun cloning in Streptococcus lactis. FEMS Microbiol. Lett. 166:355-360.
Margolin, W. 2000. Themes and variations in prokaryotic cell division. FEMS Microbiol. Rev. 24:531-548.
Marrec-Fairley, M., A. Piette, X. Gallet, R. Brasseur, H. Hara, C. Fraipont, J. M. Ghuysen, and M. Nguyen-Distèche. 2000. Differential functionalities of amphiphilic peptide segments of the cell-septation penicillin-binding protein 3 of Escherichia coli. Mol. Microbiol. 37:1019-1031.
Mercer, K. L., and D. S. Weiss. 2002. The Escherichia coli cell division protein FtsW is required to recruit its cognate transpeptidase, FtsI (PBP3), to the division site. J. Bacteriol. 184:904-912.
Mollerach, M. E., P. Partoune, J. Coyette, and J.-M. Ghuysen. 1996. Importance of the E-46-D-160 polypeptide segment of the non-penicillin-binding module for the folding of the low-affinity, multimodular class B penicillin-binding protein 5 of Enterococcus hirae. J. Bacteriol. 178:1774-1775.
Morlot, C., M. Noirclerc-Savoye, A. Zapun, O. Dideberg, and T. Vernet. 2004. The DD-carboxypeptidase PBP3 organizes the division process of Streptococcus pneumoniae. Mol. Microbiol. 51:1641-1648.
Nanninga, N. 1998. Morphogenesis of Escherichia coli. Microbiol. Mol. Biol. Rev. 62:110-129.
Nilsen, T., A. S. Ghosh, M. B. Goldberg, and K. D. Young. 2004. Branching sites and morphological abnormalities behave as ectopic poles in shape-defective Escherichia coli. Mol. Microbiol. 52:1045-1054.
Partoune, P. 1999. PhD thesis. University of Liege, Liege, Belgium.
Pastoret, S., C. Fraipont, T. Den Blaauwen, B. Wolf, M. E. G. Aarsman, A. Piette, A. Thomas, R. Brasseur, and M. Nguyen-Distèche. 2004. Functional analysis of the cell division FtsW of Escherichia coli. J. Bacteriol. 186:8370-8379.
Pavan, S., P. Hols, J. Delcour, M. C. Geoffroy, C. Grangette, M. Kleerebezem, and A. Mercenier. 2000. Adaptation of the nisin-controlled expression system in Lactobacillus plantarum: a tool to study in vivo biological effects. Appl. Environ. Microbiol. 66:4427-4432.
Piette, A., C. Fraipont, T. den Blaauwen, M. E. G. Aarsman, S. Pastoret, and M. Nguyen-Distèche. 2004. Structural determinants required to target penicillin-binding protein 3 to the septum of Escherichia coli. J. Bacteriol. 186:6110-6117.
Pinho, M. G., and J. Errington. 2003. Dispersed mode of Staphylococcus aureus cell wall synthesis in the absence of the division machinery. Mol. Microbiol. 50:871-881.
Pinho, M. G., S. R. Filipe, H. de Lancaster, and A. Tomasz. 2001. Complementation of the essential peptidoglycan transpeptidase function of penicillin-binding protein 2 (PBP2) by the drug resistance protein PBP2a in Staphylococcus aureus. J. Bacteriol. 183:6525-6531.
Popham, D. L., and K. D. Young. 2003. Role of penicillin-binding proteins in bacterial cell morphogenesis. Curr. Opin. Microbiol. 6:594-599.
Sapunaric, F., C. Franssen, P. Stefanic, A. Amoroso, O. Dardenne, and J. Coyette. 2003. Redefining the role of psr in β-lactam resistance and cell autolysis of Enterococcus hirae. J. Bacteriol. 185:5925-5935.
Satta, G., R. Fontana, and P. Canepari. 1994. The two-competing site (TCS) model of cell shape regulation in bacteria: the envelope as an integration point for the regulatory circuits of essential physiological events. Adv. Microb. Phys. 36:181-245.
Sauvage, E., F. Kerff, E. Fonzé, R. Herman, B. Schoot, J. P. Marquette, Y. Taburet, D. Prevost, J. Dumas, G. Léonard, P. Stefanic, J. Coyette, and P. Charlier. 2002. The 2.4-Å crystal structure of the penicillin-resistant penicillin-binding protein PBP5fm from Enterococcus faecium in complex with benzylpenicillin. Cell. Mol. Life Sci. 59:1223-1232.
Scheffers, D. J., and M. G. Pinho. 2005. Bacterial cell wall synthesis: new insights from localization studies. Microbiol. Mol. Biol. Rev. 69:585-607.
Sifaoui, F., M. Arthur, L. Rice, and L. Gutmann. 2001. Role of penicillin-binding protein 5 in expression of ampicillin resistance and peptidoglycan structure in Enterococcus faecium. Antimicrob. Agents Chemother. 45:2594-2597.
Signoretto, C., M. Boaretti, and P. Canepari. 1998. Peptidoglycan synthesis by Enterococcus faecalis penicillin-binding protein 5. Arch. Microbiol. 170:185-190.
Simon, D., and A. Chopin. 1988. Construction of a vector plasmid family and its use for molecular cloning in Streptococcus lactis. Biochimie 70:559-566.
Simon, M. J., and R. A. Day. 2000. Improved resolution of hydrophobic penicillin-binding proteins and their covalently linked complexes on a modified C18 reversed phase column. Anal. Lett. 33:861-867.
Spratt, B. G. 1975. Distinct penicillin-binding proteins involved in the division elongation and shape of Escherichia coli K12. Proc. Natl. Acad. Sci. USA 72:2999-3003.
Tamaki, S., H. Matsuzawa, and M. Matsuhashi. 1980. Clusters of mrdA and mrdB genes responsible for the rod shape and mecillinam sensitivity of Escherichia coli. J. Bacteriol. 141:52-57.
van Heijenoort, J. 2001. Formation of the glycan chains in the synthesis of bacterial peptidoglycan. Glycobiology 11:25R-30R.
Vollmer, W., and J. V. Höltje. 2001. Morphogenesis of Escherichia coli. Curr. Opin. Microbiol. 4:625-633.
Weiss, D. S., K. Pogliano, M. Carson, L. M. Guzman, C. Fraipont, M. Nguyen-Distèche, R. Losick, and J. Beckwith. 1997. Localization of the Escherichia coli cell division protein FtsI (PBP3) to the division site and cell pole. Mol. Microbiol. 25:671-681.
Williamson, R., C. Le Bouguénec, L. Gutmann, and T. Horaud. 1985. One or two low affinity penicillin-binding proteins may be responsible for the range of susceptibility of Enterococcus faecium to benzylpenicillin. J. Gen. Microbiol. 131:1933-1940.
Wissel, M. C., and D. S. Weiss. 2004. Genetic analysis of the cell division protein FtsI (PBP3): amino acid substitutions that impair septal localization of FtsI and recruitment of FtsN. J. Bacteriol. 186:490-502.
Young, K. D. 2003. Bacterial shape. Mol. Microbiol. 49:571-580.
Zorzi, W., X. Y. Zhou, O. Dardenne, J. Lamotte, D. Raze, J. Pierre, L. Gutmann, and J. Coyette. 1996. Structure of the low-affinity penicillin-binding protein 5 PBP5fm in wild-type and highly penicillin-resistant strains of Enterococcus faecium. J. Bacteriol. 178:4948-4957.
