Uhlinger D J, Tyagi S R, Lambeth J D
Department of Biochemistry, Emory University School of Medicine, Atlanta, Georgia 30322.
Biochemistry. 1995 Jan 17;34(2):524-7. doi: 10.1021/bi00002a017.
A peptide (RGVHFIF) from near the carboxyl terminus (residues 559-565) of gp91-phox, the large subunit of cytochrome b558, was previously shown to inhibit activation of the respiratory burst oxidase [Kleinberg, M. E., Malech, H. L., & Rotrosen, D. (1990) J. Biol. Chem. 265, 15577-15583]. The peptide has been proposed to compete with gp91-phox binding to p47-phox, one of the cytosolic oxidase components. In the present studies, we have used a semirecombinant system consisting of recombinant cytosolic factors (p47-phox, p67-phox, and Rac1) along with isolated plasma membrane to investigate the mechanism by which the peptide inhibits oxidase activation. In an in vitro translocation model, the peptide inhibited arachidonate-activated translocation of both p47-phox and p67-phox to the plasma membrane. The kinetic mechanism of inhibition was examined. Inhibition was noncompetitive or mixed with respect to not only Rac and p67-phox but also to p47-phox. We suggest that the peptide, rather than competing for cytochrome-p47-phox interactions, inhibits indirectly, perhaps by binding to and altering the conformation of cytochrome b558.
细胞色素b558的大亚基gp91-phox的羧基末端附近（残基559 - 565）的一种肽（RGVHFIF）先前已被证明可抑制呼吸爆发氧化酶的激活[克莱因伯格，M. E.，马莱克，H. L.，& 罗特罗森，D.（1990）《生物化学杂志》265，15577 - 15583]。有人提出该肽与gp91-phox竞争结合胞质氧化酶成分之一的p47-phox。在本研究中，我们使用了一个由重组胞质因子（p47-phox、p67-phox和Rac1）以及分离的质膜组成的半重组系统，来研究该肽抑制氧化酶激活的机制。在体外转位模型中，该肽抑制了花生四烯酸激活的p47-phox和p67-phox向质膜的转位。研究了抑制的动力学机制。抑制不仅对Rac和p67-phox是非竞争性或混合型的，对p47-phox也是如此。我们认为该肽不是通过竞争细胞色素-p47-phox相互作用来抑制，而是可能通过结合并改变细胞色素b558的构象来间接抑制。
