CLONING AND CHARACTERIZATION OF THE NADPH CYTOCHROME-P450 OXIDOREDUCTASE GENE FROM THE FILAMENTOUS FUNGUS ASPERGILLUS-NIGER

In this paper, we describe the cloning and molecular characterization of the Aspergillus niger cytochrome P450 reductase (CPR) gene, cprA, A ttempts to clone the cprA gene by heterologous hybridization technique s were unsuccessful, Using the polymerase chain reaction (PCR) with de generate primers based on conserved regions found in cpr genes from ot her organisms, we were able to isolate a fragment that contained part of the gene, With the aid of this fragment, a genomic fragment contain ing the entire coding region and 5' and 3' untranslated ends of the cp rA gene was isolated and sequenced, The cprA gene was introduced in mu ltiple copies in A, niger strain N402 using the amdS transformation sy stem, One of the resulting transformants, AB2-2, showed a 14-fold incr ease in CPR activity, indicating that the cloned cprA gene is function al. We analyzed the induction of cprA gene expression by several gener ally used cytochrome P450 inducers but did not find any induction of c prA gene expression, However, A, niger cprA gene expression could be i nduced by benzoic acid, which is the substrate of the highly inducible A, niger cytochrome P450 gene, bphA (cyp53), On the basis of a compar ison of the deduced protein sequence of the A, niger cprA gene with CP R proteins isolated from other organisms, the structure-function relat ionship of some conserved regions is discussed.