Biochemical and structural characterization of a monoethylhexyl phthalate hydrolase from Gordonia alkanivorans strain YC-RL2

Abstract

ObjectiveThis study aimed to investigate the effect of environmental factors on a monoethylhexyl phthalate hydrolase (MehpH) activity, model the 3-D structure of the enzyme and the interaction of the catalytic amino acid residues with the substrate. MethodsThe effect of environmental factors was determined by the standard enzyme assay. Primary structure analysis and 3-D model prediction were completed by DNAMAN (Version 2.1) and SWISS-MODEL server respectively and the results visualized by PyMOL software. Autodock tools, Swiss-PDB viewer and PyMOL were used to investigate the interactions between the enzyme and monoethylhexyl phthalate (MEHP). ResultsThe primary structure of this enzyme was similar to MehpH from Gordonia sp. P8219 with different optimum temperature and pH (40℃ and 8.0, respectively). The enzyme was stable in presence of organic solvents, detergents and ions. However, it was inhibited by 2 mol/L of Ni2+, Fe3+, Cu2+, Zn2+ ions, 1 mol/L phenylmethylsulfonyl fluoride (PMSF), 0.5 mol/L paraoxon, 1 mol/L phenyl glyoxal (PGO), 2 mol/L diethyl pyrocarbonate (DEPC) and 5 mol/L eserine. The pentapeptide motif GXSXG and catalytic triad HSD conserved in serine hydrolases were present in the sequence. The docking result showed that the amino residues Thr152 and Ser230 were much conserved among the hydrolases and closely associated with MEHP (5.8 A and 3.6 A respectively) and thus may play important roles in the catalytic process. However, MEHP was not very close to the catalytic triad acid residues Ser125, His291, Asp259. ConclusionThis study showed that MehpH in YC-RL2 was fairly stable in presence of organic solvents, detergents and metal ions indicating its application potential. The structural and catalytic analysis provides important information for further investigation of catalytic mechanism and enzymatic modification.