Background
Esterases and lipases hydrolyze short-chain esters and long-chain triglycerides, respectively, and therefore play essential roles in the synthesis and decomposition of ester bonds in the pharmaceutical and food industries. Many SGNH family esterases share high similarity in sequences. However, they have distinct enzymatic activities toward the same substrates. Due to a lack of structural information, the detailed catalytic mechanisms of these esterases remain barely investigated.
Results
In this study, we identified two SGNH family esterases, CrmE10 and AlinE4, from marine bacteria with significantly different preferences for pH, temperature, metal ion, and organic solvent tolerance despite high sequence similarity. The crystal structures of these two esterases, including wild type and mutants, were determined to high resolutions ranging from 1.18 Å to 2.24 Å. Both CrmE10 and AlinE4 were composed of five β-strands and nine α-helices, which formed one compact N-terminal α/β globular domain and one extended C-terminal domain. The aspartic residues (D178 in CrmE10/D162 in AlinE4) destabilized the conformations of the catalytic triad (Ser-Asp-His) in both esterases, and the metal ion Cd2+ might reduce enzymatic activity by blocking proton transfer or substrate binding. CrmE10 and AlinE4 showed distinctly different electrostatic surface potentials, despite the similar atomic architectures and a similar swap catalytic mechanism. When five negatively charged residues (Asp or Glu) were mutated to residue Lys, CrmE10 obtained elevated alkaline adaptability and significantly increased the enzymatic activity from 0 to 20% at pH 10.5. Also, CrmE10 mutants exhibited dramatic change for enzymatic properties when compared with the wide-type enzyme.
Conclusions
These findings offer a perspective for understanding the catalytic mechanism of different esterases and might facilitate the industrial biocatalytic applications.
; Xue-Wei, Xu