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Title: Structure of Bordetella pertussis peptidoglycan

Bordetella pertussis Tohama phases I and III were grown to the late-exponential phase in liquid medium containing (/sup 3/H)diaminopimelic acid and treated by a hot (96/sup 0/C) sodium dodecyl sulfate extraction procedure. Washed sodium dodecyl sulfate-insoluble residue from phases I and III consisted of complexes containing protein (ca. 40%) and peptidoglycan (60/sup 6/). Subsequent treatment with proteinase K yielded purified peptidoglycan which contained N-acetylglucosamine, N-acetylmuramic acid, alanine, glutamic acid, and diaminopimelic acid in molar ratios of 1:1:2:1:1 and <2% protein. Radiochemical analyses indicated that /sup 3/H added in diaminopimelic acid was present in peptidoglycan-protein complexes and purified peptidoglycan as diaminopimelic acid exclusively and that pertussis peptidoglycan was not O acetylated, consistent with it being degraded completely by hen egg white lysozyme. Muramidase-derived disaccharide peptide monomers and peptide-cross-linked dimers and higher oligomers were isolated by molecular-sieve chromatography; from the distribution of these peptidoglycan fragments, the extent of peptide cross-linking of both phase I and III peptidoglycan was calculated to be ca. 48%. Unambiguous determination of the structure of muramidase-derived pepidoglycan fragments by fast atom bombardment-mass spectrometry and tandem mass spectrometry indicated that the pertussis peptidoglycan monomer fraction was surprisingly homogeneous, consisting of >95% N-acetylglucosaminyl-N-acetylmuramyl-alanyl-glutamyl-diaminopimelyl-alanine.
Authors:
; ; ;
Publication Date:
OSTI Identifier:
6075681
Resource Type:
Journal Article
Resource Relation:
Journal Name: J. Bacteriol.; (United States); Journal Volume: 169:9
Research Org:
Indiana Univ. School of Medicine, Indianapolis
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; BACTERIA; GLUCOPROTEINS; STRUCTURAL CHEMICAL ANALYSIS; PROTEIN STRUCTURE; RADIOCHEMICAL ANALYSIS; TRITIUM COMPOUNDS; CARBOHYDRATES; CHEMICAL ANALYSIS; LABELLED COMPOUNDS; MICROORGANISMS; ORGANIC COMPOUNDS; PROTEINS; QUANTITATIVE CHEMICAL ANALYSIS; SACCHARIDES 550201* -- Biochemistry-- Tracer Techniques; 550701 -- Microbiology-- Tracer Techniques