Biotechnologically relevant enzymes and proteins

Applied Microbiology and Biotechnology

, Volume 97, Issue 4, pp 1625-1635

First online:

Cytochrome P450 reductase from Candida apicola: versatile redox partner for bacterial P450s

  • Marco GirhardAffiliated withInstitute of Biochemistry, Heinrich-Heine University Düsseldorf
  • , Florian TievesAffiliated withInstitute of Biochemistry, Heinrich-Heine University Düsseldorf
  • , Evelyne WeberAffiliated withInstitute of Technical Biochemistry, Universität Stuttgart
  • , Martha Sophia SmitAffiliated withDepartment of Microbial, Biochemical and Food Biotechnology, University of the Free State
  • , Vlada B. UrlacherAffiliated withInstitute of Biochemistry, Heinrich-Heine University Düsseldorf Email author 

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Abstract

Candida apicola belongs to a group of yeasts producing surface-active glycolipids consisting of sophorose and long-chain (ω)- or (ω-1)-hydroxy fatty acids. Hydroxylation of the fatty acids in this strain is likely catalyzed by cytochrome P450 monooxygenases (P450), which require reducing equivalents delivered via a cytochrome P450-diflavin reductase (CPR). We herein report cloning and characterization of the cpr gene from C. apicola ATCC 96134. The gene encoding a protein of 687 amino acids was cloned in Escherichia coli and the enzyme was expressed in functional form after truncation of its N-terminal putative membrane anchor. The truncated recombinant protein showed cytochrome c reducing activity (K M of 13.8 μM and k cat of 1,915 per minute). Furthermore, we herein demonstrate to our best knowledge for the first time the use of a eukaryotic CPR to transfer electrons to bacterial P450s (namely CYP109B1 and CYP154E1). Cloning and characterization of this CPR therefore is not only an important step in the study of the P450 systems of C. apicola, but also provides a versatile redox partner for the characterization of other bacterial P450s with appealing biotechnological potential. The GenBank accession number of the sequence described in this article is JQ015264.

Keywords

Candida apicola Cytochrome P450-diflavin reductase CPR P450 Heterologous expression