Kluetz M D, Schmidt P G
Biochemistry. 1977 Nov 29;16(24):5191-9. doi: 10.1021/bi00643a006.
Proton relaxation studies of the interactions with hog kidney diamine oxidase of water, substrate-analogue inhibitors, and product analogues indicate that the active site Cu(II) is not located near the oxidizing site of the enzyme, rather near the nonoxidized end of the binding substrate. The studies with histamine derivatives provide evidence for a concentration-dependent occupation of two sites. The site which is populated at high concentrations provides proximity of the imadazole ring nitrogen N1 to the Cu(II). Water binds at the Cu(II) of the native enzyme. However, this water is probably not involved in the hydrolysis of the enzyme-substrate imine bond to eliminate the first reaction product. O2 does not compete with H2O for a site on the Cu(II) ion. In the case of one of the probes, namely the ammonia (product) analogue dimethylamine, the validity of the protein relaxation results was verified by also observing the nitrogen (15N) relaxation rates of ammonia itself. The conclusion that the ammonium ions is not directly bonded to the active site Cu(II) is reached from both the proton and nitrogen relaxation experiments.
关于水、底物类似物抑制剂和产物类似物与猪肾二胺氧化酶相互作用的质子弛豫研究表明，活性位点的铜（II）并非位于酶的氧化位点附近，而是靠近结合底物的未氧化端。对组胺衍生物的研究为两个位点的浓度依赖性占据提供了证据。高浓度时占据的位点使咪唑环氮N1靠近铜（II）。水结合在天然酶的铜（II）上。然而，这种水可能不参与酶 - 底物亚胺键的水解以消除第一个反应产物。氧气不会与水竞争铜（II）离子上的位点。对于其中一种探针，即氨（产物）类似物二甲胺，通过观察氨本身的氮（15N）弛豫速率也验证了蛋白质弛豫结果的有效性。质子和氮弛豫实验均得出铵离子不直接与活性位点铜（II）结合的结论。
