尼日利亚菌素介导的H⁺、K⁺和Na⁺跨囊泡膜转运：温度跃变研究

Nigericin-mediated H+, K+ and Na+ transports across vesicular membrane: T-jump studies.

作者信息

Prabhananda B S, Ugrankar M M

机构信息

Chemical Physics Group, Tata Institute of Fundamental Research, Bombay, India.

出版信息

Biochim Biophys Acta. 1991 Dec 9;1070(2):481-91. doi: 10.1016/0005-2736(91)90090-u.

DOI:10.1016/0005-2736(91)90090-u

PMID:1764460

Abstract

The decay of delta pH across vesicular membranes by nigericin-mediated H+ and metal ion (M+) transports has been studied at 25 degrees C after creating delta pH by temperature jump (T-jump). In these experiments K+ or Na+ were chosen as M+ for the compensating flux. Theoretical expressions derived to analyse these data suggest a method for estimating the intrinsic rate constants for the translocation of nig-H (k1) and for the translocation of nig-M (k2) across membrane, from the pH dependence of the delta pH decay. The following could be inferred from the analysis of data. (a) At pH approximately 7.5 and 250 mM ion concentrations, nigericin-mediated H+ and M+ transport rates are lower in a medium of K+ than in a medium of Na+, although ionophore selectivity of nigericin towards K+ is 25-45-times higher than that towards Na+. However, at lower [M+] (approximately 50 mM) the transport rates are higher in a medium of K+ than in a medium of Na+. Such behaviours can be understood with the help of parameters determined in this work. (b) The intrinsic rate constants k1 and k2 associated with the translocations of nig-H and nig-K or nig-Na across membrane are similar in magnitude. (c) At pH approximately 7.5 translocation of nig-H is the dominant rate-limiting step in a medium containing K+. In contrast with this, at this pH, translocation of nig-M is the dominant rate-limiting step when metal ion is Na+. (d)k1 approximately k2 approximately 6.10(3) s-1 could be estimated at 25 degrees C in vesicles prepared from soyabean phospholipid, and lipid mixtures of 80% phosphatidylcholine (PC) + 20% phosphatidylethanolamine and 92% PC + 8% phosphatidic acid. (e) The apparent dissociation constants of nig-M in vesicles were estimated to be approximately 1.5.10(-3) M for K+ and 6.4.10(-2) M for Na+ (at 50 mM ion concentrations) using approximately 10(-8.45) M for the apparent dissociation constant of nig-H.

摘要

在通过温度跃升（T-jump）产生跨膜pH梯度（ΔpH）后，于25℃研究了尼日利亚菌素介导的H⁺和金属离子（M⁺）跨囊泡膜转运导致的ΔpH衰减。在这些实验中，选择K⁺或Na⁺作为用于补偿通量的M⁺。为分析这些数据而推导的理论表达式表明，一种从ΔpH衰减的pH依赖性估算尼日利亚菌素-H（nig-H）跨膜转运的固有速率常数（k1）和尼日利亚菌素-M（nig-M）跨膜转运的固有速率常数（k2）的方法。从数据分析中可推断出以下几点：（a）在pH约为7.5且离子浓度为250 mM时，尽管尼日利亚菌素对K⁺的离子载体选择性比对Na⁺高25 - 45倍，但在K⁺介质中尼日利亚菌素介导的H⁺和M⁺转运速率低于Na⁺介质中的转运速率。然而，在较低的[M⁺]（约50 mM）时，K⁺介质中的转运速率高于Na⁺介质中的转运速率。借助本工作中确定的参数可以理解这种行为。（b）与nig-H和nig-K或nig-Na跨膜转运相关的固有速率常数k1和k2在大小上相似。（c）在pH约为7.5时，在含K⁺的介质中，nig-H的转运是主要的限速步骤。与此相反，在此pH下，当金属离子为Na⁺时，nig-M的转运是主要的限速步骤。（d）在由大豆磷脂、80%磷脂酰胆碱（PC） + 20%磷脂酰乙醇胺的脂质混合物以及92% PC + 8%磷脂酸制备的囊泡中，于25℃可估算出k1≈k2≈6×10³ s⁻¹。（e）使用nig-H的表观解离常数约为10⁻⁸.⁴⁵ M，估算出囊泡中nig-M对K⁺的表观解离常数约为1.5×10⁻³ M，对Na⁺的表观解离常数约为6.4×10⁻² M（在离子浓度为50 mM时）。