Falke J J, Dernburg A F, Sternberg D A, Zalkin N, Milligan D L, Koshland D E
Department of Biochemistry, University of California, Berkeley 94720.
J Biol Chem. 1988 Oct 15;263(29):14850-8.
Cysteines are substituted at six positions in the aspartate receptor, and these mutant proteins are used to investigate three major facets of receptor structure. 1) The surface of the receptor is examined through measurement of the rate constants for chemical modification of the cysteines by aqueous reagents. Different positions exhibit a range of accessibility (for example, Cys-128 most exposed, Cys-36 most buried). 2) The transmembrane structure of the receptor is determined by reaction of the cysteines with a membrane-impermeant reagent. 3) The spatial proximities in the folded structure of specific pairs of cysteines are investigated by disulfide bond formation. These studies illustrate the usefulness of site-directed sulfhydryl chemistry in the analysis of protein structure.
天冬氨酸受体的六个位置上的半胱氨酸被替换，这些突变蛋白用于研究受体结构的三个主要方面。1）通过测量水性试剂对半胱氨酸进行化学修饰的速率常数来检测受体表面。不同位置呈现出一系列可及性（例如，Cys-128最暴露，Cys-36最埋入）。2）通过半胱氨酸与一种不能透过膜的试剂反应来确定受体的跨膜结构。3）通过二硫键形成来研究特定半胱氨酸对在折叠结构中的空间接近性。这些研究说明了定点巯基化学在蛋白质结构分析中的有用性。
