Schweri M M
Division of Basic Medical Sciences, Mercer University School of Medicine, Macon, Georgia 31207.
Neuropharmacology. 1990 Oct;29(10):901-8. doi: 10.1016/0028-3908(90)90140-m.
N-Ethylmaleimide, a nonspecific protein modifier which reacts selectively with the sulfhydryl group of cysteinyl residues under controlled conditions, irreversibly inhibited the binding of [3H]threo-(+/-)-methylphenidate to a subset of stimulant binding sites in striatal tissue membranes from the rat in vitro. The inhibition was marked by a decrease in the Bmax of binding of the radiolabelled stimulant drug, while the KD remained unchanged. Pretreatment with excess unlabelled methylphenidate afforded complete protection from inactivation of the binding site by N-ethylmaleimide. Uptake of [3H]dopamine into striatal synaptosomes was likewise reduced after treatment with N-ethylmaleimide; pretreatment with large concentrations of methylphenidate provided partial protection from inactivation of transport. These findings suggest that the stimulant recognition site on the dopamine transport complex contains one or more cysteinyl residues.
N-乙基马来酰亚胺是一种非特异性蛋白质修饰剂，在可控条件下能与半胱氨酸残基的巯基选择性反应，它在体外不可逆地抑制了[3H]苏式-（±）-甲基苯丙胺与大鼠纹状体组织膜中一部分兴奋剂结合位点的结合。这种抑制表现为放射性标记兴奋剂药物结合的最大结合容量（Bmax）降低，而解离常数（KD）保持不变。用过量未标记的甲基苯丙胺预处理可完全保护结合位点不被N-乙基马来酰亚胺灭活。用N-乙基马来酰亚胺处理后，[3H]多巴胺摄取到纹状体突触体中的量同样减少；用高浓度的甲基苯丙胺预处理可部分保护转运不被灭活。这些发现表明多巴胺转运复合物上的兴奋剂识别位点含有一个或多个半胱氨酸残基。
