Comparative cross-linking and mass spectrometry of an intact F-type ATPase suggest a role for phosphorylation

被引:120
作者
Schmidt, Carla [1 ]
Zhou, Min [1 ]
Marriott, Hazel [1 ]
Morgner, Nina [1 ]
Politis, Argyris [1 ]
Robinson, Carol V. [1 ]
机构
[1] Univ Oxford, Dept Chem, Phys & Theoret Chem Lab, Oxford OX1 3QZ, England
来源
NATURE COMMUNICATIONS | 2013年 / 4卷
基金
英国惠康基金;
关键词
EPSILON-SUBUNIT; SYNTHASE; REVEALS; F-1-ATPASE; SPINACH; IDENTIFICATION; STOICHIOMETRY; CHLOROPLASTS; COMPLEXES; ALGORITHM;
D O I
10.1038/ncomms2985
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
F-type ATPases are highly conserved enzymes used primarily for the synthesis of ATP. Here we apply mass spectrometry to the F1FO-ATPase, isolated from spinach chloroplasts, and uncover multiple modifications in soluble and membrane subunits. Mass spectra of the intact ATPase define a stable lipid 'plug' in the F-O complex and reveal the stoichiometry of nucleotide binding in the F-1 head. Comparing complexes formed in solution from an untreated ATPase with one incubated with a phosphatase reveals that the dephosphorylated enzyme has reduced nucleotide occupancy and decreased stability. By contrasting chemical cross-linking of untreated and dephosphorylated forms we show that cross-links are retained between the head and base, but are significantly reduced in the head, stators and stalk. Conformational changes at the catalytic interface, evidenced by changes in cross-linking, provide a rationale for reduced nucleotide occupancy and highlight a role for phosphorylation in regulating nucleotide binding and stability of the chloroplast ATPase.
引用
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页数:11
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