Structural determination of the sheath-forming polysaccharide of Sphaerotilus montanus using thiopeptidoglycan lyase which recognizes the 1,4 linkage between α-D-GalN and β-D-GlcA

Daisuke Kashiwabara, Keiko Kondo, Ryoji Usami, Daisuke Kan, Izuru Kawamura, Yuta Kawasaki, Michio Sato, Tadashi Nittami, Ichiro Suzuki, Masato Katahira, Minoru Takeda

研究成果: Article査読

2 被引用数 (Scopus)

抄録

Sphaerotilus natans is a filamentous sheath-forming bacterium commonly found in activated sludge. Its sheath is assembled from a thiolic glycoconjugate called thiopeptidoglycan. S. montanus ATCC-BAA-2725 is a sheath-forming member of stream biofilms, and its sheath is morphologically similar to that of S. natans. However, it exhibits heat susceptibility, which distinguishes it from the S. natans sheath. In this study, chemical composition and solid-state NMR analyses suggest that the S. montanus sheath is free of cysteine, indicating that disulfide linkage is not mandatory for sheath formation. The S. montanus sheath was successfully solubilized by N-acetylation, allowing solution-state NMR analysis to determine the sugar sequence. The sheath was susceptible to thiopeptidoglycan lyase prepared from the thiopeptidoglycan-assimilating bacterium, Paenibacillus koleovorans. The reducing ends of the enzymatic digests were labeled with 4-aminobenzoic acid ethyl ester, followed by HPLC. Two derivatives were detected, and their structures were determined. We found that the sheath has no peptides and is assembled as follows: [→4)-β-D-GlcA-(1→4)-β-D-Glc-(1→3)-β-D-GalNAc-(1→4)-α-D-GalNAc-(1→4)-α-D-GalN-(1→]n (β-D-Glc and α-D-GalNAc are stoichiometrically and substoichiometrically 3-O-acetylated, respectively). Thiopeptidoglycan lyase was thus confirmed to cleave the 1,4 linkage between α-D-GalN and β-D-GlcA, regardless of the peptide moiety. Furthermore, vital fluorescent staining of the sheath demonstrated that elongation takes place at the tips, as with the S. natans sheath.

本文言語English
ページ(範囲)992-1001
ページ数10
ジャーナルInternational Journal of Biological Macromolecules
183
DOI
出版ステータスPublished - 31 7月 2021

フィンガープリント

「Structural determination of the sheath-forming polysaccharide of Sphaerotilus montanus using thiopeptidoglycan lyase which recognizes the 1,4 linkage between α-D-GalN and β-D-GlcA」の研究トピックを掘り下げます。これらがまとまってユニークなフィンガープリントを構成します。

引用スタイル