Ionization behavior of the catalytic carboxyls of lysozyme. Effects of ionic strength.
Ionization behavior of the catalytic carboxyls of lysozyme. Effects of ionic strength.
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溶菌酶催化羧基的电离行为。
DOI：
10.1021/bi00759a013
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发表时间：
1972
期刊：
影响因子：
2.9
通讯作者：
M. Raftery
中科院分区：
文献类型：
--
作者：
S. Parsons;M. Raftery
S. M.帕森斯和。A.摘要：本文研究了溶菌酶天冬氨酸52位残基的β-乙酯衍生物相对于天然溶菌酶的pH差滴定。差异曲线反映了Asp-52和Glu-35的电离行为。四个微常数描述了两个相互作用的催化基团的电离。宏观电离常数，包括所有形式的酶，为天冬氨酸-52为4.5，而谷氨酸-35在0.15 m KCl在25。每个基团的电离都强烈地依赖于离子强度，其方式与I ^相反。最近，我们制备、分离并鉴定了一种由催化残基天冬氨酸52的β-乙酯组成的鸡蛋清溶菌酶衍生物（Parsons等人，1969; Parsons and Raftery，1969）.通过对这一特性良好的衍生物和天然溶菌酶进行差分测量，我们有望获得有关活性位点区域的详细信息，这些活性位点区域的作用被蛋白质其余部分抵消了。当处理单酯化酶时，一个明显的比较是质子差滴定。值得注意的是，这是一个优选的组合，即使另一种方法，如pH依赖性扰动蛋白紫外（uv）吸光度是可用的。这是因为氢离子滴定具有可预测的化学计量范围。因此，可以置信地拟合更复杂的差异曲线。
S. M. ParsonsJ and. A. Raftery* abstract: The pH difference titration of the/3-ethyl ester derivative of the Asp-52 residue of lysozyme relative to native lysozyme has been obtained. The difference curve reflects the ionization behavior of both Asp-52 and Glu-35. Four micro-constants describe the ionizations of the two interacting cata-lytic groups. The macroscopic ionization constant, including all forms of the enzyme, for Asp-52 is 4.5 while that for Glu-35 is 5.9 in 0.15 m KC1 at 25. The ionization of each group is strongly dependent on ionic strength in a manner opposite toI^. ecently we prepared, isolated, and identified a deriva-tive of hen egg-white lysozyme which consists of the/3-ethyl ester of the catalytic residue aspartic acid 52 (Parsons et al., 1969; Parsons and Raftery, 1969). By subjecting this wellcharacterized derivative and native lysozyme to a differential measurement we could hope to gain detailed knowledge about the active site regionin which effects from the rest of the protein were cancelled out. An obvious comparison to make when dealing with a singly esterified enzyme is a proton differ-ence titration. It is worth noting that this is a preferred com-parison even if another method such as pH-dependent per-turbed protein ultraviolet (uv) absorbance is available. This is so because thehydrogen ion titration has a predictable stoi-chiometric span. Accordingly, more complex difference curves can be fitted with confidence.