Cloning and expression of a cDNA encoding the β-subunit (30-kDa subunit) of bovine brain platelet-activating factor acetylhydrolase

Mitsuharu Hattori, Hideki Adachi, Junken Aoki, Masafumi Tsujimoto, Hiroyuki Arai, Keizo Inoue

研究成果: Article査読

85 被引用数 (Scopus)

抄録

Bovine brain platelet-activating factor (PAF) acetylhydrolase isoform Ib is a heterotrimeric enzyme. Its γ-subunit (which, formerly, we called the 29-kDa subunit) acts as a catalytic subunit, whereas the α-subunit (45 kDa) is the bovine homolog of the product of human LIS-1, the causative gene of Miller-Dieker lissencephaly, indicating that this intracellular PAF acetylhydrolase plays a key role in brain development. In the current study, we cloned the cDNA for the β-subunit (30 kDa) of bovine brain PAF acetylhydrolase Ib. The predicted 229-amino acid sequence was homologous (63.2% identity) to that of the γ-subunit, especially (86% identity) in the catalytic and PAF receptor homologous domains. The recombinant β-protein produced in Escherichia coli showed significant PAF acetylhydrolase activity. A mutant protein, in which Ser48, which corresponds to the active serine residue of the γ-subunit, was replaced with cysteine showed no enzymatic activity, suggesting Ser48 is the active serine residue. Although the β- and γ-subunits form a heterocomplex in the native enzyme, both recombinant β- and γ-proteins exist as a homodimer. The purified recombinant β- protein was labeled readily with [1,3-3H]diisopropyl fluorophosphate, whereas the β-subunit in the native complex was only labeled with higher concentrations of [1,33H]diisopropyl fluorophosphate to a lesser extent than the γ-subunit. Combined with our previous data, the present study demonstrated that bovine brain PAF acetylhydrolase Ib is a unique enzyme possessing two catalytic subunits and another, possibly regulatory, subunit.

本文言語English
ページ(範囲)31345-31352
ページ数8
ジャーナルJournal of Biological Chemistry
270
52
DOI
出版ステータスPublished - 1995 12 29

ASJC Scopus subject areas

  • 生化学
  • 分子生物学
  • 細胞生物学

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