期刊
BIOCHIMICA ET BIOPHYSICA ACTA-BIOENERGETICS
卷 1777, 期 1, 页码 32-38出版社
ELSEVIER SCIENCE BV
DOI: 10.1016/j.bbabio.2007.11.004
关键词
ATP synthase; FIFO; subunit a; proton translocation; ATP synthesis; rotary motor
资金
- NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES [R01GM040508] Funding Source: NIH RePORTER
- NIGMS NIH HHS [R01 GM040508, GM40508, R01 GM040508-18] Funding Source: Medline
Interactions between subunit a and oligomeric subunit e are essential for the coupling of proton translocation to rotary motion in the ATP synthase. A pair of previously described mutants, R210Q/Q252R and P204T/R210Q/Q252R [L.P. Hatch, G.B. Cox and S.M. Howitt, The essential arginine residue at position 210 in the a subunit of the Escherichia coli ATP synthase can be transferred to position 252 with partial retention of activity, J. Biol. Chem. 270 (1995) 29407-29412] has been constructed and further analyzed. These mutants, in which the essential arginine of subunit a, R210, was switched with a conserved glutamine residue, Q252, are shown here to be capable of both ATP synthesis by oxidative phosphorylation, and ATP-driven proton translocation. In addition, lysine can replace the arginine at position 252 with partial retention of both activities. The pH dependence of ATP-driven proton translocation was determined after purification of mutant enzymes, and reconstitution into liposomes. Proton translocation by the lysine mutant, and to a lesser extent the arginine mutant, dropped off sharply above pH 7.5, consistent with the requirement for a positive charge during function. Finally, the rates of ATP synthesis and of ATP-driven proton translocation were completely inhibited by treatment with DCCD (N,N'-dicyclohexylcarbodiimide), while rates of ATP hydrolysis by the mutants were not significantly affected, indicating that DCCD modification disrupts the F-I-F-O interface. The results suggest that minimal requirements for proton translocation by the ATP synthase include a positive charge in subunit a and a weak interface between subunit a and oligomeric subunit c. (C) 2007 Elsevier B.V. All rights reserved.
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