PURIFICATION AND CHARACTERIZATION OF A HIGHLY SELECTIVE EPOXIDE HYDROLASE FROM NOCARDIA SP. EH1

A highly enantioselective, soluble epoxide hydrolase from Nocardia sp. EH1 was purified to homogeneity via a four-step procedure: (i) hydrop hobic interaction chromatography on Phenyl Sepharose CL-4B, (ii) anion exchange chromatography on SOURCE 30Q, followed by (iii) a second hyd rophobic interaction chromatography on Phenyl Sepharose HP, and finall y (iv) gel-filtration on Superdex 75 HR 10/30. The pure protein was sh own to be a monomer of similar to 34 kDa possessing an optimum pH of 8 -9. Neither UV-absorbing cofactors nor metal ions were required for ac tivity. In contrast to whole-cell activity, the partially purified enz yme proved to be considerably less stable. Stabilization was achieved by addition of non-ionic detergents such as Tween 80 or Triton X-100, causing a shift of the temperature optimum from 35 to 40 degrees C. Bo th effects combined led to an enhancement of the relative activity of up to similar to 150% of that of the native enzyme. (C) 1998 Elsevier Science B.V. All rights reserved.