Lyophilization conditions for the storage of monooxygenases

J Biotechnol. 2015 Jun 10:203:41-4. doi: 10.1016/j.jbiotec.2015.03.010. Epub 2015 Mar 26.

Abstract

Cyclohexanone monooxygenase (CHMO) was used as a model enzyme to find suitable freeze-drying conditions for long-term storage of an isolated monooxygenase. CHMO is a Baeyer-Villiger monooxygenase (BVMO) known for its ability to catalyze a large number of oxidation reactions. With a focus on establishing the optimal formulation, additives were tested for enzyme stabilization during and after lyophilization. The results were successfully transferred to two other monooxygenases, namely the BVMO cyclopentadecanone monooxygenase (CPDMO) and a cytochrome P450 monooxygenase, P450 BM3. In the absence of a lyoprotectant, lyophilized P450 BM3 is almost completely inactivated, while the lyophilized BVMOs quickly lost activity when stored at 50°C. Lyophilization in the presence of 2% (w/v) sucrose was found to be the best formulation to preserve activity and protect against inactivation when stored as lyophilizate at 50°C.

Keywords: Biocatalyst; Cryoprotection; Freeze-drying; Lyophilization; Monooxygenase; Sucrose.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacterial Proteins / chemistry*
  • Cytochrome P-450 Enzyme System / chemistry*
  • Drug Storage
  • Freeze Drying
  • NADPH-Ferrihemoprotein Reductase / chemistry*
  • Oxygenases / chemistry*

Substances

  • Bacterial Proteins
  • Cytochrome P-450 Enzyme System
  • Oxygenases
  • cyclohexanone oxygenase
  • NADPH-Ferrihemoprotein Reductase
  • flavocytochrome P450 BM3 monoxygenases