Binding of inhibitory metals to yeast enolase.
Binding of inhibitory metals to yeast enolase.
复制标题
抑制性金属与酵母烯醇酶的结合。
DOI：
10.1016/s0162-0134(00)80273-1
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发表时间：
1980
影响因子：
3.9
通讯作者：
J. Brewer
中科院分区：
文献类型：
--
作者：
J. Elliott;J. Brewer
某些二价阳离子可以通过结合在与通常与催化活性相关的那些金属结合位点不同的位点来抑制酵母烯醇化酶，即，构象和催化结合位点。通过使用不与金属离子（四丙基硼酸铵）竞争的缓冲液，发现Zn、Co、Mn、Cu、Cd和Ni表现出类似的抑制特性。通过加入咪唑或tris缓冲液，以及对于锌，通过金属螯合剂（钙黄绿素）减轻这些金属的抑制。通过结合研究和酶活性测量详细检查了锌的抑制作用。在pH8.0的四丙基铵缓冲液中，烯醇化酶每摩尔酶结合多达4摩尔锌（每个亚基2摩尔）。0.05 M的咪唑浓度降低了结合：在没有底物的情况下，每个酶只有两摩尔的锌被结合。在任一缓冲液中加入底物后，酶将结合另外两摩尔的锌，但在四丙基铵缓冲液中的酶几乎无活性。因此，抑制作用与每摩尔酶结合两摩尔锌有关。一些额外的金属离子，钙，铽，汞，和银也引起酵母烯醇化酶的抑制，但不是通过结合到所述的抑制位点。
Certain divalent cations can inhibit yeast enolase by binding at sites that are distinct from those metal binding sites normally associated with catalytic activity, i.e., the conformational and catalytic binding sites. By using a buffer that does not compete with metal ions (tetrapropylammonium borate) Zn, Co, Mn, Cu, Cd, and Ni are found to exhibit similar inhibitory characteristics. Inhibition by those metals is alleviated by the addition of imidazole or tris buffer and, for zinc, by a metal chelating agent (Calcein). Inhibition by zinc was examined in detail through binding studies and enzymatic activity measurement. In tetrapropylammonium buffers at pH 8.0, enolase binds up to four moles of zinc per mole of enzyme (two moles per subunit). An imidazole concentration of 0.05 M reduces the binding: in the absence of substrate, just two moles of zinc per enzyme are bound. The enzyme will bind two additional moles of zinc upon the addition of substrate in either buffer, but the enzyme in tetrapropylammonium buffer is nearly inactive. Inhibition is, therefore, correlated with the binding of two moles of zinc per mole of enzyme. Some additional metal ions, Ca, Tb, Hg, and Ag also caused inhibition of yeast enolase but not by binding to the inhibitory site described.