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Publication Abstract from PubMed

To investigate the mechanism for low-pH adaptation by a carboxylesterase, structural and biochemical analyses of EstFa_R (a recombinant, slightly acidophilic carboxylesterase from Ferroplasma acidiphilum) and SshEstI (an alkaliphilic carboxylesterase from Sulfolobus shibatae DSM5389) were performed. Although a previous proteomic 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 ICP-MS and isothermal titration calorimetry. The crystal structures of EstFa_R and SshEstI were determined at 1.6A and 1.5A 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., 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 negative 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.

Structural Insights into the Low-pH Adaptation of a Unique Carboxylesterase from Ferroplasma: Altering the pH Optima of Two Carboxylesterases.,Ohara K, Unno H, Oshima Y, Hosoya M, Fujino N, Hirooka K, Takahashi S, Yamashita S, Kusunoki M, Nakayama T J Biol Chem. 2014 Jul 20. pii: jbc.M113.521856. PMID:25043762^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.