cDNA cloning and expression of the mRNA for cytochrome P‐450kd which shows a fatty acid ω‐hydroxylating activity

Noboru YOKOTANI, Emi KUSUNOSE, Kazuhiro SOGAWA, Hidenori KAWASHIMA, Masahiko KINOSAKI, Masamichi KUSUNOSE, Yoshiaki FUJII‐KURIYAMA

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    30 Citations (Scopus)

    Abstract

    We have recently purified three distinct forms of fatty acid ω‐hydroxylase cytochrome P‐450 (P‐450), designated P‐450ka‐1, P‐450ka‐2 and P‐450kd, from rabbit kidney cortex microsomes, and isolated and sequenced cDNA clones corresponding to P‐450ka‐1 and P‐450ka‐2 [Yokotani, N., Bernhardt, R., Sogawa, K., Kusunose, E., Gotoh, M., Kusunose, M. & Fujii‐Kuriyama, Y. (1989) J. Biol. Chem. 264, 21665–21669]. The present paper describes cloning, sequencing and expression of a cDNA for the third fatty acid, ω‐hydroxylase, P‐450kd, from a rabbit kidney cDNA library. The cDNA for P‐450kd encodes a polypeptide of 511 amino acids with sequence similarity of 87% to P‐450ka‐1. Its deduced NH2‐terminal sequence of amino acids 5–24 is in complete agreement with the NH2‐terminal sequence of P‐450kd. The identity of the cDNA was further confirmed by its expression in COS‐7 cells. When 14C‐labeled lauric acid was added to the culture medium of COS‐7 cells transfected with the cDNA, significant amounts of radioactive dodecanedioic acid, together with ω‐ and (ω‐1)‐hydroxylauric acids, were produced. Microsomes prepared from the transfected cells also efficiently catalyzed the ω‐ and (ω‐1)‐hydroxylation of lauric acid without formation of dodecanedioic acid. RNA blot analysis demonstrated that the mRNA for P‐450kd gave a single band at the approximately 2.6‐kb position. The mRNA for P‐450kd was expressed in the liver and kidney, but not in many other tissues examined. Treatment of rabbits with clofibrate resulted in a elevated level of mRNA for P‐450kd in both liver and kidney. Furthermore, the mRNA was remarkably increased in the kidney by the administration of cyclosporin A.

    Original languageEnglish
    Pages (from-to)531-536
    Number of pages6
    JournalEuropean Journal of Biochemistry
    Volume196
    Issue number3
    DOIs
    Publication statusPublished - 1991 Mar

    ASJC Scopus subject areas

    • Biochemistry

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