[en] BACKGROUND: Proteins from the LytR-CpsA-Psr family are found in almost all Gram-positive bacteria. Although LCP proteins have been studied in other pathogens, their functions in enterococci remain uncharacterized. The Psr protein from Enterococcus hirae, here renamed LcpA, previously associated with the regulation of the expression of the low-affinity PBP5 and β-lactam resistance, has been characterized.
RESULTS: LcpA protein of E. hirae ATCC 9790 has been produced and purified with and without its transmembrane helix. LcpA appears, through different methods, to be localized in the membrane, in agreement with in silico predictions. The interaction of LcpA with E. hirae cell wall indicates that LcpA binds enterococcal peptidoglycan, regardless of the presence of secondary cell wall polymers. Immunolocalization experiments showed that LcpA and PBP5 are localized at the division site of E. hirae.
CONCLUSIONS: LcpA belongs to the LytR-CpsA-Psr family. Its topology, localization and binding to peptidoglycan support, together with previous observations on defective mutants, that LcpA plays a role related to the cell wall metabolism, probably acting as a phosphotransferase catalyzing the attachment of cell wall polymers to the peptidoglycan.
Disciplines :
Biochemistry, biophysics & molecular biology
Author, co-author :
Maréchal, Maxime ; Université de Liège - ULiège > Centres généraux > Centre d'ingénierie des protéines
Amoroso, Ana Maria ; Université de Liège - ULiège > Département des sciences de la vie > Centre d'Ingénierie des Protéines (CIP)
Morlot, Cécile; University Grenoble Alpes, IBS, Grenoble, F-38044, France ; CNRS, IBS, Grenoble, F-38044, France ; CEA, IBS, Grenoble, F-38044, France
Vernet, Thierry; University Grenoble Alpes, IBS, Grenoble, F-38044, France ; CNRS, IBS, Grenoble, F-38044, France ; CEA, IBS, Grenoble, F-38044, France
Coyette, Jacques ; Université de Liège - ULiège > Département des sciences de la vie > Physiologie et génétique bactériennes
Joris, Bernard ; Université de Liège - ULiège > Département des sciences de la vie > Centre d'Ingénierie des Protéines (CIP)
Language :
English
Title :
Enterococcus hirae LcpA (Psr), a new peptidoglycan-binding protein localized at the division site.
F.R.S.-FNRS - Fonds de la Recherche Scientifique [BE] ANR - Agence Nationale de la Recherche [FR] Grenoble Alpes University [FR] FRIA - Fonds pour la Formation à la Recherche dans l'Industrie et dans l'Agriculture [BE]
Funding number :
University Grenoble Alpes (Fond d’intervention CSVSB 2011); Belgian Federal Government (IAP program P7/44 iPROS)
Funding text :
Support for this work comes in part from the Belgian Federal Government (IAP program P7/44 iPROS), the FRS-FNRS Fonds de la Recherche Scientifique, CC-7057031, the Agence Nationale de la Recherche (ANR-11-BSV8-005-01 PILIPATH) and the University Grenoble Alpes (Fond d’intervention CSVSB 2011). This work used the platforms of the Grenoble Instruct center (ISBG; UMS 3518 CNRS-CEA-UJF-EMBL) with support from FRISBI (grant ANR-10-INSB-05-02) and GRAL (grant ANR-10-LABX-49-01) within the Grenoble Partnership for Structural Biology. MM was recipient of a fellowship from the Fonds pour la Recherche dans l’Industrie et l’Agriculture (FRIA).
Murray BE. The life and times of the Enterococcus. Clin Microbiol Rev. 1990;3(1):46-65.
Bhavsar AP, Brown ED. Cell wall assembly in Bacillus subtilis: how spirals and spaces challenge paradigms. Mol Microbiol. 2006;60(5):1077-90.
Vollmer W, Blanot D, de Pedro MA. Peptidoglycan structure and architecture. FEMS Microbiol Rev. 2008;32(2):149-67.
Hakenbeck R, Coyette J. Resistant penicillin-binding proteins. Cell Mol Life Sci. 1998;54(4):332-40.
Zapun A, Contreras-Martel C, Vernet T. Penicillin-binding proteins and beta-lactam resistance. FEMS Microbiol Rev. 2008;32(2):361-85.
Klein G. Taxonomy, ecology and antibiotic resistance of enterococci from food and the gastro-intestinal tract. Int J Food Microbiol. 2003;88(2-3):123-31.
Fontana R, Cerini R, Longoni P, Grossato A, Canepari P. Identification of a streptococcal penicillin-binding protein that reacts very slowly with penicillin. J Bacteriol. 1983;155(3):1343-50.
Canepari P, Lleo MM, Cornaglia G, Fontana R, Satta G. In Streptococcus faecium penicillin-binding protein 5 alone is sufficient for growth at sub-maximal but not at maximal rate. J Gen Microbiol. 1986;132(3):625-31.
Sifaoui F, Arthur M, Rice L, Gutmann L. Role of penicillin-binding protein 5 in expression of ampicillin Resistance and peptidoglycan structure in enterococcus faecium. Antimicrob Agents Chemother. 2001;45(9):2594-7.
Leimanis S, Hoyez N, Hubert S, Laschet M, Sauvage E, Brasseur R, Coyette J. PBP5 complementation of a PBP3 deficiency in Enterococcus hirae. J Bacteriol. 2006;188(17):6298-307.
Rice LB, Carias LL, Rudin S, Hutton R, Marshall S, Hassan M, Josseaume N, Dubost L, Marie A, Arthur M. Role of class A penicillin-binding proteins in the expression of beta-lactam resistance in Enterococcus faecium. J Bacteriol. 2009;191(11):3649-56.
Gaechter T, Wunderlin C, Schmidheini T, Solioz M. Genome sequence of Enterococcus hirae (Streptococcus faecalis) ATCC 9790, a model organism for the study of ion transport, bioenergetics, and copper homeostasis. J Bacteriol. 2012;194(18):5126-7.
Hancock LE, Murray BE, Sillanpää J. Enterococcal Cell Wall Components and Structures, Enterococci: From Commensals to Leading Causes of Drug Resistant Infection. 2014.
Hancock LE, Gilmore MS. The capsular polysaccharide of Enterococcus faecalis and its relationship to other polysaccharides in the cell wall. Proc Natl Acad Sci U S A. 2002;99(3):1574-9.
Teng F, Singh KV, Bourgogne A, Zeng J, Murray BE. Further characterization of the epa gene cluster and Epa polysaccharides of Enterococcus faecalis. Infect Immun. 2009;77(9):3759-67.
Ligozzi M, Pittaluga F, Fontana R. Identification of a genetic element (psr) which negatively controls expression of Enterococcus hirae penicillin-binding protein 5. J Bacteriol. 1993;175(7):2046-51.
Shockman GD, Daneo-Moore L, Kariyama R, Massidda O. Bacterial walls, peptidoglycan hydrolases, autolysins, and autolysis. Microb Drug Resist. 1996;2(1):95-8.
Daneo-Moore L, Massidda O, Kariyama R, Shockman GD. Penicillin resistance and autolysis in enterococci. Microb Drug Resist. 1996;2(1):159-61.
Massidda O, Kariyama R, Daneo-Moore L, Shockman GD. Evidence that the PBP5 synthesis repressor (psr) of Enterococcus hirae is also involved in the regulation of cell wall composition and other cell wall-related properties. J Bacteriol. 1996;178(17):5272-8.
Rice LB, Carias LL, Hutton-Thomas R, Sifaoui F, Gutmann L, Rudin SD. Penicillin-binding protein 5 and expression of ampicillin resistance in Enterococcus faecium. Antimicrob Agents Chemother. 2001;45(5):1480-6.
Duez C, Zorzi W, Sapunaric F, Amoroso A, Thamm I, Coyette J. The penicillin resistance of Enterococcus faecalis JH2-2r results from an overproduction of the low affinity penicillin-binding protein PBP4 and does not involve a psr-like gene. Microbiology. 2001;147(9):2561-9.
Sapunaric F, Franssen C, Stefanic P, Amoroso A, Dardenne O, Coyette J. Redefining the role of psr in beta-lactam Resistance and cell autolysis of Enterococcus hirae. J Bacteriol. 2003;185(20):5925-35.
Massidda O, Dardenne O, Whalen M, Zorzi W, Coyette J, Shockman GD, Daneo-Moore L. The PBP 5 synthesis repressor (psr) gene of Enterococcus hirae ATCC 9790 is substantially longer than previously reported. FEMS Microbiol Lett. 1998;166(2):355-60.
Hubscher J, Luthy L, Berger-Bachi B, Stutzmann Meier P. Phylogenetic distribution and membrane topology of the LytR-CpsA-Psr protein family. BMC Genomics. 2008;9:617.
Over B, Heusser R, McCallum N, Schulthess B, Kupferschmied P, Gaiani JM, Sifri CD, Berger-Bachi B, Stutzmann Meier P. LytR-CpsA-Psr proteins in Staphylococcus aureus display partial functional redundancy and the deletion of all three severely impairs septum placement and cell separation. FEMS Microbiol Lett. 2011;320(2):142-51.
Kawai Y, Marles-Wright J, Cleverley RM, Emmins R, Ishikawa S, Kuwano M, Heinz N, Bui NK, Hoyland CN, Ogasawara N, et al. A widespread family of bacterial cell wall assembly proteins. EMBO J. 2011;30(24):4931-41.
Bitoun JP, Liao S, McKey BA, Yao X, Fan Y, Abranches J, Beatty WL, Wen ZT. Psr is involved in regulation of glucan production, and double deficiency of BrpA and Psr is lethal in Streptococcus mutans. Microbiology. 2013;159(Pt 3):493-506.
Eberhardt A, Hoyland CN, Vollmer D, Bisle S, Cleverley RM, Johnsborg O, Havarstein LS, Lewis RJ, Vollmer W. Attachment of capsular polysaccharide to the cell wall in Streptococcus pneumoniae. Microb Drug Resist. 2012;18(3):240-55.
Dengler V, Meier PS, Heusser R, Kupferschmied P, Fazekas J, Friebe S, Staufer SB, Majcherczyk PA, Moreillon P, Berger-Bachi B, et al. Deletion of hypothetical wall teichoic acid ligases in Staphylococcus aureus activates the cell wall stress response. FEMS Microbiol Lett. 2012;333(2):109-20.
Chan YG, Frankel MB, Dengler V, Schneewind O, Missiakas D. Staphylococcus aureus mutants lacking the LytR-CpsA-Psr family of enzymes release cell wall teichoic acids into the extracellular medium. J Bacteriol. 2013;195(20):4650-9.
Chan YG, Kim HK, Schneewind O, Missiakas D. The capsular polysaccharide of Staphylococcus aureus is attached to peptidoglycan by the LytR-CpsA-Psr (LCP) family of enzymes. J Biol Chem. 2014;289(22):15680-90.
Wu C, Huang IH, Chang C, Reardon-Robinson ME, Das A, Ton-That H. Lethality of sortase depletion in Actinomyces oris caused by excessive membrane accumulation of a surface glycoprotein. Mol Microbiol. 2014;94(6):1227-41.
Amer BR, Clubb RT. A sweet new role for LCP enzymes in protein glycosylation. Mol Microbiol. 2014;94(6):1197-200.
Liszewski Zilla M, Chan YG, Lunderberg JM, Schneewind O, Missiakas D. LytR-CpsA-Psr enzymes as determinants of Bacillus anthracis secondary cell wall polysaccharide assembly. J Bacteriol. 2015;197(2):343-53.
Liszewski Zilla M, Lunderberg JM, Schneewind O, Missiakas D. Bacillus anthracis lcp genes support vegetative growth, envelope assembly, and spore formation. J Bacteriol. 2015;197(23):3731-41.
Hanson BR, Runft DL, Streeter C, Kumar A, Carion TW, Neely MN. Functional analysis of the CpsA protein of Streptococcus agalactiae. J Bacteriol. 2012;194(7):1668-78.
Rowe HM, Hanson BR, Runft DL, Lin Q, Firestine SM, Neely MN. Modification of the CpsA protein reveals a role in alteration of the Streptococcus agalactiae cell envelope. Infect Immun. 2015;83(4):1497-506.
Wang Q, Zhu L, Jones V, Wang C, Hua Y, Shi X, Feng X, Jackson M, Niu C, Gao Q. CpsA, a LytR-CpsA-Psr family protein in mycobacterium marinum, Is required for cell Wall integrity and virulence. Infect Immun. 2015;83(7):2844-54.
Mistou MY, Sutcliffe IC, van Sorge NM. Bacterial glycobiology: rhamnose-containing cell wall polysaccharides in Gram-positive bacteria, FEMS microbiology reviews. 2016.
Chahboune A, Decaffmeyer M, Brasseur R, Joris B. Membrane topology of the Escherichia coli AmpG permease required for recycling of cell wall anhydromuropeptides and AmpC beta-lactamase induction. Antimicrob Agents Chemother. 2005;49(3):1145-9.
Broome-Smith JK, Tadayyon M, Zhang Y. Beta-lactamase as a probe of membrane protein assembly and protein export. Mol Microbiol. 1990;4(10):1637-44.
Morlot C, Zapun A, Dideberg O, Vernet T. Growth and division of Streptococcus pneumoniae: localization of the high molecular weight penicillin-binding proteins during the cell cycle. Mol Microbiol. 2003;50(3):845-55.
Daniel RA, Errington J. Control of cell morphogenesis in bacteria. Cell. 2003;113(6):767-76.
Tiyanont K, Doan T, Lazarus MB, Fang X, Rudner DZ, Walker S. Imaging peptidoglycan biosynthesis in Bacillus subtilis with fluorescent antibiotics. Proc Natl Acad Sci U S A. 2006;103(29):11033-8.
Girardin SE, Boneca IG, Carneiro LA, Antignac A, Jehanno M, Viala J, Tedin K, Taha MK, Labigne A, Zahringer U, et al. Nod1 detects a unique muropeptide from gram-negative bacterial peptidoglycan. Science. 2003;300(5625):1584-7.
Kerff F, Amoroso A, Herman R, Sauvage E, Petrella S, Filee P, Charlier P, Joris B, Tabuchi A, Nikolaidis N, et al. Crystal structure and activity of Bacillus subtilis YoaJ (EXLX1), a bacterial expansin that promotes root colonization. Proc Natl Acad Sci U S A. 2008;105(44):16876-81.
Manoil C, Beckwith J. A genetic approach to analyzing membrane protein topology. Science. 1986;233(4771):1403-8.
