Leukotriene A4 hydrolase, a bifunctional enzyme Distinction of leukotriene A4 hydrolase and aminopeptidase activities by site‐directed mutagenesis at Glu‐297

FEBS Letters

Tsuge, Hideaki³  Shimada, Shinobu²  Izumi, Takashi²  Minami, Michiko²  Abe, Keiko¹  Ohishi, Nobuya²  Mutoh, Hiroyuki²  Wada, Hiroo²  Bito, Haruhiko²  Miyano, Masashi³  Mori, Mikiro²  Shimizu, Takao² 

[1] Department of Agricultural Chemistry, Faculty of Agriculture, University of Tokyo, Bunkyo-ku, Tokyo 113, Japan;Department of Biochemistry, Faculty of Medicine, University of Tokyo, Bunkyo-ku, Tokyo 113, Japan;Life Science Research Laboratory Japan Tabacco Inc., 6-2 Umegaoka, Midoriku, Yokohama, Kanagawa, Japan

关键词: Leukotriene;    Leukotriene A4 hydrolase;    Aminopeptidase;    Zinc-metalloprotease;    Site-directed mutagenesis;   

DOI  :  10.1016/0014-5793(92)80806-R

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

We previously obtained evidence for intrinsic aminopeptidase activity for leukotriene (LT)A₄ hydrolase, an enzyme characterized to specifically catalyse the hydrolysis of LTA₄ to LTB₄, a chemotactic compound. From a sequence homology search between LTA₄ hydrolase and several aminopeptidases, it became clear that they share a putative active site for known aminopeptidases and a zinc binding domain. Thus, Glu-297 of LTA₄ hydrolase is a candidate for the active site of its aminopeptidase activity, while His-296, His-300 and Glu-319 appear to constitute a zinc binding site. To determine whether or not this putative active site is also essential to LTA₄ hydrolase activity, site-directed mutagenesis experiments were carried out. Glu-297 was mutated into 4 different amino acids. The mutant E297Q (Glu changed to Gln) conserved LTA₄ hydrolase activity but showed little aminopeptidase activity. Other mutants at Glu-297 (E297A, E297D and E297K) showed markedly reduced amounts of both activities. It is thus proposed that either a glutamic or glutamine moiety at 297 is required for full LTA₄ hydrolase activity, while the free carboxylic acid of glutamic acid is essential for aminopeptidase.

【 授权许可】

Unknown