Elliott J I, Brewer J M
J Inorg Biochem. 1980 Jul;12(4):323-34. doi: 10.1016/s0162-0134(00)80273-1.
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 zine 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.
某些二价阳离子可通过结合在与通常与催化活性相关的金属结合位点不同的位点（即构象和催化结合位点）来抑制酵母烯醇化酶。通过使用不与金属离子竞争的缓冲液（四丙基硼酸铵），发现锌、钴、锰、铜、镉和镍表现出相似的抑制特性。这些金属的抑制作用可通过添加咪唑或三羟甲基氨基甲烷缓冲液来缓解，对于锌，可通过金属螯合剂（钙黄绿素）来缓解。通过结合研究和酶活性测量详细研究了锌的抑制作用。在pH 8.0的四丙基铵缓冲液中，烯醇化酶每摩尔酶可结合多达四摩尔锌（每个亚基两摩尔）。0.05 M的咪唑浓度会降低结合：在没有底物的情况下，每个酶仅结合两摩尔锌。在任何一种缓冲液中添加底物后，该酶将再结合两摩尔锌，但四丙基铵缓冲液中的酶几乎没有活性。因此，抑制作用与每摩尔酶结合两摩尔锌相关。一些其他金属离子，如钙、铽、汞和银也会抑制酵母烯醇化酶，但不是通过结合到所述的抑制位点来实现的。
