单价阳离子诱导的亚线粒体颗粒内囊泡pH变化：与Na⁺/H⁺和K⁺/H⁺反向转运体的关系

Intravesicular pH changes in submitochondrial particles induced by monovalent cations: relationship to the Na+/H+ and K+/H+ antiporters.

作者信息

Brierley G P, Davis M H, Jung D W

机构信息

Department of Physiological Chemistry, Ohio State University Medical Center, Columbus 43210.

出版信息

Arch Biochem Biophys. 1988 Aug 1;264(2):417-27. doi: 10.1016/0003-9861(88)90307-4.

DOI:10.1016/0003-9861(88)90307-4

PMID:2456741

Abstract

The fluorescence of internalized fluorescein isothiocyanate dextran has been used to monitor the intravesicular pH of submitochondrial particles (SMP). Respiring SMP maintain a steady-state delta pH (interior acid) that results from the inwardly directed H+ flux of respiration and an opposing passive H+ leak. Addition of K+, Na+, or Li+ to SMP results in a shift to a more alkaline interior pH (pHi) in both respiring and nonrespiring SMP. The K+-dependent change in pHi, like the K+/H+ antiport in intact mitochondria, is inhibited by quinine and by dicyclohexylcarbodiimide. The Na+-dependent reaction is only partially inhibited by these reagents. Both the Na+- and the K+-dependent pH changes are sensitive to amiloride derivatives. The Km for both Na+ and K+ is near 20 mM whereas that for Li+ is closer to 10 mM. The K+/H+ exchange reaction is only slightly inhibited by added Mg2+, but abolished when A23187 is added with Mg2+. The passive exchange is optimal at pHi 6.5 with either Na+ or K+, and cannot be detected above pHi of 7.2. Both the Na+/H+ and the K+/H+ exchange reactions are optimal at an external pH of 7.8 in respiring SMP (pHi 7.1). Valinomycin stimulates the K+-dependent pH change in nonrespiring SMP, as does nigericin. It is concluded that SMP show K+/H+ antiport activity with properties distinct from those of Na+/H+ antiport. However, the properties of the K+/H+ exchange do not correspond in all respects to those of the antiport in intact mitochondria. Donnan equilibria and parallel uniport pathways for H+ and cations appear to contribute to cation-dependent pH changes in SMP.

摘要

内化的异硫氰酸荧光素葡聚糖的荧光已被用于监测亚线粒体颗粒（SMP）的囊泡内pH值。进行呼吸作用的SMP维持着一个稳态的ΔpH（内部呈酸性），这是由呼吸作用中向内的H⁺通量和相反方向的被动H⁺泄漏导致的。向SMP中添加K⁺、Na⁺或Li⁺会导致进行呼吸作用和不进行呼吸作用的SMP的内部pH（pHi）都向更碱性的方向转变。pHi的K⁺依赖性变化，就像完整线粒体中的K⁺/H⁺反向转运一样，受到奎宁和二环己基碳二亚胺的抑制。Na⁺依赖性反应仅被这些试剂部分抑制。Na⁺和K⁺依赖性的pH变化都对氨氯地平衍生物敏感。Na⁺和K⁺的Km值接近20 mM，而Li⁺的Km值更接近10 mM。K⁺/H⁺交换反应仅被添加的Mg²⁺轻微抑制，但当A23187与Mg²⁺一起添加时则被消除。被动交换在pHi为6.5时，无论是对于Na⁺还是K⁺都是最佳的，并且在pHi高于7.2时无法检测到。在进行呼吸作用的SMP（pHi 7.1）中，Na⁺/H⁺和K⁺/H⁺交换反应在外部pH为7.8时都是最佳的。缬氨霉素刺激不进行呼吸作用的SMP中K⁺依赖性的pH变化，尼日利亚菌素也有同样的作用。得出的结论是，SMP表现出K⁺/H⁺反向转运活性，其特性与Na⁺/H⁺反向转运不同。然而，K⁺/H⁺交换的特性在所有方面并不都与完整线粒体中的反向转运特性一致。唐南平衡以及H⁺和阳离子的平行单向转运途径似乎导致了SMP中阳离子依赖性的pH变化。