Analyses of the cell-wall peptidoglycan structures in three genera Micromonospora, Catenuloplanes, and Couchioplanes belonging to the family Micromonosporaceae by derivatization with FDLA and PMP using LC/MS

Akira Take, Takuji Nakashima, Yuki Inahashi, Kazuro Shiomi, Yōko Takahashi, Satoshi Ōmura, Atsuko Matsumoto

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Abstract

It is the major characteristic of the cell-wall peptidoglycan structure in members of the family Micromonosporaceae that N-acetylmuramic acid (MurNAc) of glycan strand is replaced with Nglycolylmuramic acid (MurNGlyc). Consequently, it is difficult to use enzymatic methods for their peptidoglycan analyses. We therefore developed analysis method of peptidoglycan without using cell wall lytic enzymes as example to take the 3 genera, Micromonospora, Catenuloplanes, and Couchioplanes belonging to the family Micromonosporaceae, and their peptidoglycans were partially hydrolyzed with 4 M HCl at 60°C for 16 h followed by derivatization with Nα-(5- fluoro-2,4-dinitrophenyl)-D-leucinamide (FDLA) or 1-phenyl-3-methyl-5-pyrazolone (PMP) and LC/ MS analysis. Peptidoglycan of the genus Micromonospora consisted of a MurNGlyc–Gly–DGlu–meso-diaminopimelyl (DAP)–D-Ala peptide stem and direct linkage between D-Ala and meso- DAP. In contrast, peptidoglycans of the genera Catenuloplanes and Couchioplanes consisted of a MurNGlyc–Gly–D-Glu–L-Lys–D-Ala peptide stem, and cross-linkage between D-Ala and L-Lys was mediated by an L-Ser residue. This method can be used to analyze the cell-wall peptidoglycan structure of other bacteria as well. By derivatization with FDLA or PMP followed by LC/MS analysis, the structure can be determined using only 0.2 mg of purified peptidoglycan.

Original languageEnglish
Pages (from-to)199-205
Number of pages7
JournalJournal of General and Applied Microbiology
Volume62
Issue number4
DOIs
Publication statusPublished - 2016 Jan 1
Externally publishedYes

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Keywords

  • Catenuloplanes
  • Couchioplanes
  • FDLA
  • LC/MS
  • Micromonospora
  • Peptidoglycan
  • PMP

ASJC Scopus subject areas

  • Microbiology
  • Applied Microbiology and Biotechnology

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