title: Subcellular localization of the major autolysin, ATL and its processed proteins in Staphylococcus aureus.
abstract: The Staphylococcus aureus autolysin gene, atl, encodes a unique 138-kDa protein (ATL) with amidase and glucosaminidase domains. ATL has been suggested to undergo proteolytic processing to generate two extracellular peptidoglycan hydrolases, 51-kDa endo-beta-N-acetylglucosaminidase (51-kDa GL) and 62-kDa N-acetylmuramyl-L-alanine amidase (62-kDa AM). To investigate cell-associated bacteriolytic enzymes for atl gene products, proteins were extracted from the cells as follows. The cells were exposed to 3 M LiCl followed by 4% SDS. Thereafter, the cells were disrupted and again extracted with 4% SDS. Whole SDS-stable cell-associated bacteriolytic proteins were extracted without disrupting the cells. Exposure to 3 M LiCl released major 138-, 115-, 85-, 62- and 51-kDa bacteriolytic proteins, and subsequent 4% SDS extraction released major 138- and 115-kDa bacteriolytic proteins. These bacteriolytic proteins were missing in extracts of atl mutant RUSAL2 (S. aureus RN450 atl::Tn551). Immunoblotting studies suggest that these are all atl gene products: the 138-kDa protein is an ATL with a cleaved signal sequence; the 115- and 85-kDa proteins are intermediates; and the 51- and 62-kDa proteins are cell-associated 51-kDa GL and 62-kDa AM, respectively. The trypsin susceptibility of these proteins suggests that they are located outside the cell membrane. Differences in extractability and immunoelectron microscopic studies suggest that atl gene products are associated with cells in two different ways, LiCl extractable and non extractable. We suggest that the 138-kDa ATL undergoes processing through intermediate proteins (115- and 85-kDa proteins) to mature as the active cell cluster-dispersing enzymes 51-kDa GL and 62-kDa AM on the cell surface.
authors.affiliation: Department of Microbiology, Hiroshima University School of Dentistry, Japan.
authors.forename: H
authors.initials: H
authors.lastname: Komatsuzawa
authors.forename: M
authors.initials: M
authors.lastname: Sugai
authors.forename: S
authors.initials: S
authors.lastname: Nakashima
authors.forename: S
authors.initials: S
authors.lastname: Yamada
authors.forename: A
authors.initials: A
authors.lastname: Matsumoto
authors.forename: T
authors.initials: T
authors.lastname: Oshida
authors.forename: H
authors.initials: H
authors.lastname: Suginaka
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification*; D001433:Bacteriolysis; D004792:Enzyme Precursors / Q000235:genetics / Q000302:isolation & purification*; D018021:Lithium Chloride / Q000494:pharmacology; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase / Q000235:genetics; D008969:Molecular Sequence Data; D009113:Muramidase / Q000235:genetics; D009154:Mutation; D009238:N-Acetylmuramoyl-L-alanine Amidase / Q000235:genetics / Q000302:isolation & purification*; D011489:Protein Denaturation; D011499:Protein Processing, Post-Translational; D013211:Staphylococcus aureus / Q000201:enzymology* / Q000378:metabolism / Q000648:ultrastructure
chemical_list: D001426:Bacterial Proteins; D004792:Enzyme Precursors; D009113:Muramidase; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D009238:N-Acetylmuramoyl-L-alanine Amidase; D018021:Lithium Chloride