Effects of denaturants on amide proton exchange rates: a test for structure in protein fragments and folding intermediates.
Effects of denaturants on amide proton exchange rates: a test for structure in protein fragments and folding intermediates.
复制标题
变性剂对酰胺质子交换率的影响：蛋白质片段和折叠中间体结构的测试。
DOI：
10.1021/bi00354a036
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发表时间：
1986
期刊：
影响因子：
2.9
通讯作者：
Baldwin,RL
中科院分区：
文献类型：
--
作者：
Loftus,D;Gbenle,GO;Kim,PS;Baldwin,RL
接收于1985年7月25日的摘要：描述了一种检测蛋白质边缘稳定形式结构的方法。原理是测量酰胺质子交换率在不存在和存在不同浓度的变性剂。在对变性剂对固有交换速率的影响进行校正后，变性剂对结构的展开反映在酰胺质子交换速率的加速上。这种结构的汇率检验没有对未展开状态下的汇率作任何假设。标定了0-8 M尿素和0-6 M氯化胍（GdmCl）对模型化合物酸碱催化交换的影响。GdmCl似乎不太适合用于汇率测试；模型复合研究表明，国内生产总值对内在汇率的影响是复杂的。相比之下，尿素的作用是变性剂浓度的更均匀的函数。尿素增加了模型化合物酸催化的交换率，降低了碱催化的交换率。这里使用汇率试验来研究s蛋白（核糖核酸酶A残基21-124）的结构形成。在已知s蛋白（I3）的平衡折叠中间体填充的条件下（pH为1.7,0 C），结构的交换率测试是阳性的。在较高温度（bbb32℃）下，I3展开，但圆二色性数据表明残余结构仍然存在[Labhardt， AM (1982) J. Mol. Biol. 157, 357-371]。在这些条件下（pH 1.7, 45℃），交换率测试为负，表明热展开s蛋白的任何残余结构对酰胺质子交换率没有可检测到的影响。
Department of Biochemistry, Stanford University, Stanford, California 94305 Received July 25, 1985 abstract: A method for detecting structure in marginally stable forms of a protein is described. The principle is to measure amide proton exchange rates in the absence and presence of varying concentrations of a denaturant. Unfolding of structure by the denaturant is reflected by an acceleration of amide proton exchange rates, after correction for theeffects of the denaturant on the intrinsic rate of exchange. This exchange-rate test for structure makes no assumptions about the rate of exchange in the unfolded state. The effects of 0-8 M urea and 0-6 M guanidinium chloride (GdmCl) on acid-and base-catalyzed exchange from model compounds havebeen calibrated. GdmCl does not appear tobe well-suited for use in the exchange-rate test; model compound studies show that theeffects of GdmCl on intrinsic exchange rates are complicated. In contrast, the effects of urea are a more uniform function of denaturant concentration. Urea increases acid-catalyzed, anddecreases base-catalyzed, exchange rates in model compounds. The exchange-rate test is used here to study structure formation in the S-protein (residues 21-124 of ribonuclease A). In conditions where an equilibrium folding intermediate of S-protein (I3) is known to be populated (pH 1.7, 0 C), the exchange-rate test for structure is positive. At higher temperatures (> 32 C) I3 is unfolded, but circular dichroism data suggest that residual structure remains [Labhardt, AM (1982) J. Mol. Biol. 157, 357-371], Under these conditions (pH 1.7, 45 C) the exchange-rate test is negative, indicating that any residual structure in thermally unfolded S-protein does not have a detectable effect on amide proton exchange rates.