Structure of Bordetella pertussis peptidoglycan.
Bordetella pertussis Tohama phases I and III were grown to the late-exponential phase in liquid medium containing [3H]diaminopimelic acid and treated by a hot (96 degrees 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%). 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 less than 2% protein. Radiochemical analyses indicated that 3H 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 peptidoglycan fragments by fast atom bombardment-mass spectrometry and tandem mass spectrometry indicated that the pertussis peptidoglycan monomer fraction was surprisingly homogeneous, consisting of greater than 95% N-acetylglucosaminyl-N-acetylmuramyl-alanyl-glutamyl-diaminopimelyl++ +-alanine.
D000107:Acetylation; D000596:Amino Acids / Q000032:analysis; D001886:Bordetella pertussis / Q000032:analysis* / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D002854:Chromatography, Paper; D003960:Diaminopimelic Acid / Q000032:analysis / Q000378:metabolism; D013058:Mass Spectrometry; D009113:Muramidase / Q000378:metabolism; D010457:Peptidoglycan / Q000032:analysis*
D000596:Amino Acids; D010457:Peptidoglycan; D003960:Diaminopimelic Acid; D009113:Muramidase