title: Substrate recognition and catalysis by LytB, a pneumococcal peptidoglycan hydrolase involved in virulence.
abstract: Streptococcus pneumoniae is a major cause of life-threatening diseases worldwide. Here we provide an in-depth functional characterization of LytB, the peptidoglycan hydrolase responsible for physical separation of daughter cells. Identified herein as an N-acetylglucosaminidase, LytB is involved also in colonization and invasion of the nasopharynx, biofilm formation and evasion of host immunity as previously demonstrated. We have shown that LytB cleaves the GlcNAc-β-(1,4)-MurNAc glycosidic bond of peptidoglycan building units. The hydrolysis occurs at sites with fully acetylated GlcNAc moieties, with preference for uncross-linked muropeptides. The necessity of GlcN acetylation and the presence of a single acidic moiety (Glu585) essential for catalysis strongly suggest a substrate-assisted mechanism with anchimeric assistance of the acetamido group of GlcNAc moieties. Additionally, modelling of the catalytic region bound to a hexasaccharide tripentapeptide provided insights into substrate-binding subsites and peptidoglycan recognition. Besides, cell-wall digestion products and solubilisation rates might indicate a tight control of LytB activity to prevent unrestrained breakdown of the cell wall. Choline-independent localization at the poles of the cell, mediated by the choline-binding domain, peptidoglycan modification, and choline-mediated (lipo)teichoic-acid attachment contribute to the high selectivity of LytB. Moreover, so far unknown chitin hydrolase and glycosyltransferase activities were detected using GlcNAc oligomers as substrate.
authors.affiliation: Instituto de Química-Física Rocasolano, Consejo Superior de Investigaciones Científicas, Serrano 119, 28006, Madrid, Spain.|CIBER de Enfermedades Respiratorias (CIBERES), Instituto de Salud Carlos III (ISCIII), Madrid, Spain.
authors.forename: Palma
authors.initials: P
authors.lastname: Rico-Lastres
authors.affiliation: CIBER de Enfermedades Respiratorias (CIBERES), Instituto de Salud Carlos III (ISCIII), Madrid, Spain.|Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Ramiro de Maeztu 9, 28040, Madrid, Spain.
authors.forename: Roberto
authors.initials: R
authors.lastname: Díez-Martínez
authors.affiliation: Instituto de Química-Física Rocasolano, Consejo Superior de Investigaciones Científicas, Serrano 119, 28006, Madrid, Spain.|CIBER de Enfermedades Respiratorias (CIBERES), Instituto de Salud Carlos III (ISCIII), Madrid, Spain.
authors.forename: Manuel
authors.initials: M
authors.lastname: Iglesias-Bexiga
authors.affiliation: Instituto de Química-Física Rocasolano, Consejo Superior de Investigaciones Científicas, Serrano 119, 28006, Madrid, Spain.|CIBER de Enfermedades Respiratorias (CIBERES), Instituto de Salud Carlos III (ISCIII), Madrid, Spain.
authors.forename: Noemí
authors.initials: N
authors.lastname: Bustamante
authors.affiliation: Centre for Bacterial Cell Biology, Institute for Cell and Molecular Biosciences, Newcastle University, Newcastle upon Tyne NE2 4AX, United Kingdom.
authors.forename: Christine
authors.initials: C
authors.lastname: Aldridge
authors.affiliation: Department of Chemistry and Biochemistry, University Notre Dame, Notre Dame, Indiana 46556, United States.
authors.forename: Dusan
authors.initials: D
authors.lastname: Hesek
authors.affiliation: Department of Chemistry and Biochemistry, University Notre Dame, Notre Dame, Indiana 46556, United States.
authors.forename: Mijoon
authors.initials: M
authors.lastname: Lee
authors.affiliation: Department of Chemistry and Biochemistry, University Notre Dame, Notre Dame, Indiana 46556, United States.
authors.forename: Shahriar
authors.initials: S
authors.lastname: Mobashery
authors.affiliation: Institute for Cell and Molecular Biosciences, Newcastle University, Newcastle upon Tyne NE2 4AX, United Kingdom.
authors.forename: Joe
authors.initials: J
authors.lastname: Gray
authors.affiliation: Centre for Bacterial Cell Biology, Institute for Cell and Molecular Biosciences, Newcastle University, Newcastle upon Tyne NE2 4AX, United Kingdom.
authors.forename: Waldemar
authors.initials: W
authors.lastname: Vollmer
authors.affiliation: CIBER de Enfermedades Respiratorias (CIBERES), Instituto de Salud Carlos III (ISCIII), Madrid, Spain.|Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Ramiro de Maeztu 9, 28040, Madrid, Spain.
authors.forename: Pedro
authors.initials: P
authors.lastname: García
authors.affiliation: Instituto de Química-Física Rocasolano, Consejo Superior de Investigaciones Científicas, Serrano 119, 28006, Madrid, Spain.|CIBER de Enfermedades Respiratorias (CIBERES), Instituto de Salud Carlos III (ISCIII), Madrid, Spain.
authors.forename: Margarita
authors.initials: M
authors.lastname: Menéndez
mesh_terms: D000118:Acetylglucosaminidase / Q000378:metabolism; D002384:Catalysis; D020134:Catalytic Domain / Q000502:physiology; D002473:Cell Wall / Q000378:metabolism; D002686:Chitin / Q000378:metabolism; D002794:Choline / Q000378:metabolism; D016695:Glycosyltransferases / Q000378:metabolism; D006867:Hydrolases / Q000378:metabolism; D006868:Hydrolysis; D009238:N-Acetylmuramoyl-L-alanine Amidase / Q000378:metabolism*; D009305:Nasopharynx / Q000382:microbiology; D010457:Peptidoglycan / Q000378:metabolism*; D013296:Streptococcus pneumoniae / Q000378:metabolism* / Q000472:pathogenicity*; D013379:Substrate Specificity; D013682:Teichoic Acids / Q000378:metabolism; D014774:Virulence
chemical_list: D010457:Peptidoglycan; D013682:Teichoic Acids; D002686:Chitin; D016695:Glycosyltransferases; D006867:Hydrolases; D000118:Acetylglucosaminidase; D009238:N-Acetylmuramoyl-L-alanine Amidase; D002794:Choline