Role of the gamma subunit of chloroplast coupling factor 1 in the light-dependent activation of photophosphorylation and ATPase activity by dithiothreitol.
Role of the gamma subunit of chloroplast coupling factor 1 in the light-dependent activation of photophosphorylation and ATPase activity by dithiothreitol.
复制标题
DOI：
10.1016/s0021-9258(17)39870-8
复制
发表时间：
1984-06
期刊：
影响因子：
--
通讯作者：
Stuart R. Ketcham;James W. Davenport;Kurt WarnckelI;R. E. Mccarty
中科院分区：
文献类型：
--
作者：
Stuart R. Ketcham;James W. Davenport;Kurt WarnckelI;R. E. Mccarty
在叶片和完整的叶绿体中，氧化和还原已被证明调节类囊体的ATPase活性。在二硫苏糖醇存在下，菠菜叶绿体类囊体光照能激活类囊体体在黑暗中催化持续的ATP水解的能力，导致N-乙基马来酰亚胺掺入偶联因子1(CF1)的伽马亚基。在活化过程中，伽马亚基中的二硫键被还原。这个二硫键所涉及的残基与二硫苏糖醇活化可溶性CF1时还原的二硫键中的残基相同。在十二烷基硫酸钠存在下，可以用聚丙烯酰胺凝胶电泳法分离伽马亚基的二硫键和二硫醇形式。N-乙基马来酰亚胺优先在黑暗中掺入先前暴露于二硫苏糖醇的类囊体伽马亚基CF1的还原形式中。在黑暗中，类囊体中只有CF1的一部分对二硫苏糖醇还原和随后与N-乙基马来酰亚胺的反应敏感。通过还原二硫键暴露的硫醇基团的烷基化可以保护ATPase活性免受氧化剂的抑制。在一个给定的跨膜pH差值下，二硫苏糖醇激活的类囊体光磷酸化速率可以是非激活对照的七到八倍。N-乙基马来酰亚胺在黑暗中处理激活的类囊体能防止失去对类囊体储存时的ATP合成的刺激。与洗涤的类囊体相比，在测定混合物中裂解的完整叶绿体的光磷酸化也被激活。在较低的ADP浓度下，随着三角洲pH的升高，光合磷酸化速率接近饱和。这些结果表明，CF1的伽马亚基在调节ATP合成和水解酶方面起着重要作用。
In leaves and intact chloroplasts, oxidation and reduction have been shown previously to regulate the ATPase activity of thylakoids. Illumination of spinach chloroplast thylakoids in the presence of dithiothreitol, which activates the ability of thylakoids to catalyze sustained ATP hydrolysis in the dark, causes increased incorporation of N-ethylmaleimide into the gamma subunit of coupling factor 1 (CF1). A disulfide bond in the gamma subunit is reduced during activation. The residues involved in this disulfide bond are the same as those in the disulfide linkage reduced during dithiothreitol activation of soluble CF1. The disulfide and dithiol forms of the gamma subunit may be separated by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. N-Ethylmaleimide is preferentially incorporated in the dark into the reduced form of the gamma subunit of CF1 in thylakoids previously exposed to dithiothreitol. Only a subpopulation of the CF1 in thylakoids is susceptible to dithiothreitol reduction and subsequent reaction with N-ethylmaleimide in the dark. Alkylation of the thiol groups exposed by reduction of the disulfide bond protects ATPase activity from inhibition by oxidants. At a given value of the transmembrane pH differential, photophosphorylation rates in dithiothreitol-activated thylakoids can be as much as seven to eight times those of nonactivated controls. N-Ethylmaleimide treatment of activated thylakoids in the dark prevents the loss of the stimulation of ATP synthesis on storage of the thylakoids. Photophosphorylation by intact chloroplasts lysed in assay mixtures is also activated in comparison to that by washed thylakoids. At a low ADP concentration, the rate of photophosphorylation approaches saturation as delta pH increases. These results suggest that the gamma subunit of CF1 plays an important role in regulation of ATP synthesis and hydrolysis.