Cloning and expression of Koala )Phascolarctos cinerus) liver cytochrome P450 reductase

S KONG, Suong Ngo, R MCKINNON, L STUPANS

    Research output: Contribution to journalArticle

    Abstract

    The cloning, expression and characterization of hepatic NADPH-cytochrome P450 reductase (CPR) from koala (Phascolarctos cinereus) is described. Two 2059�bp koala liver CPR cDNAs, designated CPR1 and CPR2, were cloned by reverse transcription-polymerase chain reaction and rapid amplification of cDNA ends. The koala CPR cDNAs encode proteins of 678 amino acids and share 85% amino acid sequence identity to human CPR. Transfection of the koala CPR cDNAs into Cos-7 cells resulted in the expression of proteins, which were recognized by a goat-antihuman CPR antibody. The koala CPR1 and 2 cDNA-expressed enzymes catalysed cytochrome c reductase at the rates of 4.9 � 0.5 and 2.6 � 0.4�nmol/min/mg protein (mean � SD, n = 3), respectively which were comparable to that of rat CPR cDNA-expressed enzyme. The apparent Km value for CPR activity in koala liver microsomes was 11.61 � 6.01�?M, which is consistent with that reported for rat CPR enzyme. Northern analysis detected a CPR mRNA band of approximately 2.6�kb. Southern analysis suggested a single PCR gene across species. The present study provides primary molecular data regarding koala CPR1 and CPR2 genes in this unique marsupial species. � 2009 Elsevier Inc. All rights reserved.
    Original languageEnglish
    Pages (from-to)1-9
    Number of pages9
    JournalComparative Biochemistry and Physiology, Part C
    Volume150
    Issue number1
    Publication statusPublished - 2009

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