Molecular Cloning and Crystal Structural Analysis of a Novel β-N-Acetylhexosaminidase from Paenibacillus sp. TS12 Capable of Degrading Glycosphingolipids

Tomomi Sumida, Ryohei Ishii, Tatsuo Yanagisawa, Shigeyuki Yokoyama, Makoto Ito

研究成果: ジャーナルへの寄稿学術誌査読

39 被引用数 (Scopus)

抄録

We report the molecular cloning and characterization of two novel β-N-acetylhexosaminidases (β-HEX, EC 3.2.1.52) from Paenibacillus sp. strain TS12. The two β-HEXs (Hex1 and Hex2) were 70% identical in primary structure, and the N-terminal region of both enzymes showed significant similarity with β-HEXs belonging to glycoside hydrolase family 20 (GH20). Interestingly, however, the C-terminal region of Hex1 and Hex2 shared no sequence similarity with the GH20 β-HEXs or other known proteins. Both recombinant enzymes, expressed in Escherichia coli BL21(DE3), hydrolyzed the β-N-acetylhexosamine linkage of chitooligosaccharides and glycosphingolipids such as asialo GM2 and Gb4Cer in the absence of detergent. However, the enzyme was not able to hydrolyze GM2 ganglioside in the presence or in the absence of detergent. We determined three crystal structures of Hex1; the Hex1 deletion mutant Hex1-ΔC at a resolution of 1.8 Å; Hex1-ΔC in complex with β-N-acetylglucosamine at 1.6 Å; and Hex1-ΔC in complex with β-N-acetylgalactosamine at 1.9 Å. We made a docking model of Hex1-ΔC with GM2 oligosaccharide, revealing that the sialic acid residue of GM2 could hinder access of the substrate to the active site cavity. This is the first report describing the molecular cloning, characterization and X-ray structure of a procaryotic β-HEX capable of hydrolyzing glycosphingolipids.

本文言語英語
ページ(範囲)87-99
ページ数13
ジャーナルJournal of Molecular Biology
392
1
DOI
出版ステータス出版済み - 9月 11 2009

!!!All Science Journal Classification (ASJC) codes

  • 構造生物学
  • 分子生物学

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