Duque R E, Phan S H, Sulavik M C, Ward P A
J Biol Chem. 1983 Jul 10;258(13):8123-8.
A plasma membrane-associated serine esterase (protease) has previously been reported to be required during stimulation of a variety of cell types. In this study we have examined the role of such a protease on membrane potential changes using an optical probe of membrane potential 3,3'-dipropylthiodicarbocyanine iodide. The irreversible chymotrypsin-like protease inhibitor TPCK (tosyl-L-phenylalanyl chloromethyl ketone) inhibited membrane potential changes in rat neutrophils in response to phorbol myristate acetate, N-formylmethionylleucylphenylalanine and the calcium ionophore A23187 in a time- and dose-dependent manner. This inhibition was correlated with the known inhibitory effects on superoxide (O-2) generation and enzyme release (lysozyme and beta-glucuronidase). High external calcium concentrations overcame the inhibitory effects of TPCK on A23187-induced stimulation but had no effect on the inhibition of N-formylmethionylleucylphenylalanine stimulation. These results suggest that a TPCK-inhibitable activity is required for the development of the membrane potential changes that are coupled to subsequent secretory events and precede a calcium requiring activity.
先前有报道称，在多种细胞类型的刺激过程中需要一种与质膜相关的丝氨酸酯酶（蛋白酶）。在本研究中，我们使用膜电位光学探针3,3'-二丙基硫代二碳菁碘化物，研究了这种蛋白酶在膜电位变化中的作用。不可逆的类胰凝乳蛋白酶抑制剂TPCK（甲苯磺酰-L-苯丙氨酰氯甲基酮）以时间和剂量依赖性方式抑制大鼠中性粒细胞对佛波酯、N-甲酰甲硫氨酰亮氨酰苯丙氨酸和钙离子载体A23187的膜电位变化。这种抑制作用与对超氧化物（O-2）生成和酶释放（溶菌酶和β-葡萄糖醛酸酶）的已知抑制作用相关。高细胞外钙浓度克服了TPCK对A23187诱导刺激的抑制作用，但对N-甲酰甲硫氨酰亮氨酰苯丙氨酸刺激的抑制作用没有影响。这些结果表明，一种可被TPCK抑制的活性是与随后的分泌事件相关且先于钙依赖性活性的膜电位变化发展所必需的。
