Structural insights into the low pH adaptation of a unique carboxylesterase from Ferroplasma: Altering the pH optima of two carboxylesterases

Kazuhiro Ohara, Hideaki Unno, Yasuhiro Oshima, Miho Hosoya, Naoto Fujino, Kazutake Hirooka, Seiji Takahashi, Satoshi Yamashita, Masami Kusunoki, Toru Nakayama

研究成果: Article査読

23 被引用数 (Scopus)


To investigate the mechanism for low pH adaptation by a carboxylesterase, structural and biochemical analyses of Est-Fa-R (a recombinant, slightly acidophilic carboxylesterase from Ferroplasma acidiphilum ) and SshEstI (an alkaliphilic carboxylesterase from Sulfolobus shibatae DSM5389) were performed. Although a previous proteomics study by another group showed that the enzyme purified from F. acidiphilum contained an iron atom, EstFa-R did not bind to iron as analyzed by inductively coupled plasma MS and isothermal titration calorimetry. The crystal structures of EstFa-R and SshEstI were determined at 1.6- and 1.5-Å resolutions, respectively. EstFa-R had a catalytic triad with an extended hydrogen bond network that was not observed in SshEstI. Quadruple mutants of both proteins were created to remove or introduce the extended hydrogen bond network. The mutation on EstFa-R enhanced its catalytic efficiency and gave it an alkaline pH optimum, whereas the mutation on SshEstI resulted in opposite effects ( i.e. a decrease in the catalytic efficiency and a downward shift in the optimum pH). Our experimental results suggest that the low pH optimum of EstFa-R activity was a result of the unique extended hydrogen bond network in the catalytic triad and the highly negatively charged surface around the active site. The change in the pH optimum of EstFa-R happened simultaneously with a change in the catalytic efficiency, suggesting that the local flexibility of the active site in EstFa-R could be modified by quadruple mutation. These observations may provide a novel strategy to elucidate the low pH adaptation of serine hydrolases.

ジャーナルJournal of Biological Chemistry
出版ステータスPublished - 2014

ASJC Scopus subject areas

  • 生化学
  • 分子生物学
  • 細胞生物学


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