Module shuffling of a family F/10 xylanase: Replacement of modules M4 and M5 of the FXYN of Streptomyces olivaceoviridis E-86 with those of the Cex of Cellulomonas fimi

Satoshi Kaneko*, Shinnosuke Iwamatsu, Atsushi Kuno, Zui Fujimoto, Yoko Sato, Kei Yura, Mitiko Go, Hiroshi Mizuno, Kazunari Taira, Tsunemi Hasegawa, Isao Kusakabe, Kiyoshi Hayashi

*この研究の対応する著者

研究成果: Article査読

19 被引用数 (Scopus)

抄録

To facilitate an understanding of structure-function relationships, chimeric xylanases were constructed by module shuffling between the catalytic domains of the FXYN from Streptomyces olivaceoviridis E-86 and the Cex from Cellulomonas fimi. In the family F/10 xylanases, the modules M4 and M5 relate to substrate binding so that modules M4 and M5 of the FXYN were replaced with those of the Cex and the chimeric enzymes denoted FCF-C4, FCF-C5 and FCF-C4,5 were constructed. The kcat value of FCF-C5 for p-nitrophenyl-β-D-cellobioside was similar to that of the FXYN (2.2 s-1); however, the kcat value of FCF-C4 for p-nitrophenyl-β-D-cellobioside was significantly higher (7.0 s-1). The loss of the hydrogen bond between E46 and S22 or the presence of the I49W mutation would be expected to change the position of Q88, which plays a pivotal role in discriminating between glucose and xylose, resulting in the increased kcat value observed for FCF-C4 acting on p-nitrophenyl-β-D-cellobioside since module M4 directly interacts with Q88. To investigate the synergistic effects of the different modules, module M10 of the FCF-C4 chimera was replaced with that of the Cex. The effects of replacement of module M4 and M10 were almost additive with regard to the Km and kcat values.

本文言語English
ページ(範囲)873-879
ページ数7
ジャーナルProtein Engineering
13
12
DOI
出版ステータスPublished - 2000
外部発表はい

ASJC Scopus subject areas

  • 生化学
  • 分子生物学

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