Modi S
Department of Biochemistry, University of Cambridge, UK.
Biochim Biophys Acta. 1993 Mar 5;1162(1-2):121-6. doi: 10.1016/0167-4838(93)90137-g.
Interaction of EDTA with horseradish peroxidase (HRP) was investigated by NMR relaxation-rate measurements. At pH 4.0, the apparent dissociation constant (Kd) for the EDTA binding to HRP was deduced to be 78 mM from the relaxation measurements. From pH-dependence of 1H-NMR line width of EDTA, it was observed that EDTA binds to HRP only under acidic conditions (pH < 5). The binding of EDTA to HRP was facilitated by protonation of an acid group on the enzyme with pKa 4.0. The Kd for EDTA binding to HRP was also evaluated in the presence of an excess of exogenous substrates such as iodide and thiocyanate ions. The Kd in the presence of iodide and thiocyanate ions showed that EDTA competes with these ions for the same binding site. The distance of EDTA protons from ferric centre of HRP were deduced from 1H-NMR relaxation measurements and was found to be in the order of 8 A.
通过核磁共振弛豫速率测量研究了乙二胺四乙酸（EDTA）与辣根过氧化物酶（HRP）的相互作用。在pH 4.0时，根据弛豫测量推断EDTA与HRP结合的表观解离常数（Kd）为78 mM。从EDTA的1H-NMR线宽的pH依赖性观察到，EDTA仅在酸性条件下（pH < 5）与HRP结合。EDTA与HRP的结合通过酶上pKa为4.0的酸性基团的质子化而促进。在存在过量外源性底物如碘离子和硫氰酸根离子的情况下，也评估了EDTA与HRP结合的Kd。在碘离子和硫氰酸根离子存在下的Kd表明，EDTA与这些离子竞争相同的结合位点。通过1H-NMR弛豫测量推断出EDTA质子与HRP铁中心的距离，发现其约为8埃。
