The product of uncI gene in F1Fo-ATP synthase operon plays a chaperone-like role to assist c-ring assembly
- *ATP Synthesis Regulation Project, ICORP, Japan Science and Technology Corporation, Aomi 2-41, Tokyo 135-0064, Japan; and
- †Chemical Resources Laboratory, Tokyo Institute of Technology, Nagatsuta 4259, Yokohama 226-8503, Japan
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Edited by H. Ronald Kaback, University of California, Los Angeles, CA, and approved October 30, 2007 (received for review August 27, 2007)
Abstract
Bacterial operons for F1Fo-ATP synthase typically include an uncI gene that encodes a function-unknown small hydrophobic protein. When we expressed a hybrid F1Fo (F1 from thermophilic Bacillus PS3 and Na+-translocating Fo from Propionigenium modestum) in Escherchia coli cells, we found that uncI derived from P. modestum was indispensable to produce active enzyme; without uncI, c-subunits in F1Fo existed as monomers but not as functional c 11-ring. When uncI was expressed from another plasmid at the same time, active F1Fo with c 11-ring was produced. A plasmid containing only uncI and c-subunit gene produced c 11-ring, but a plasmid containing only c-subunit gene did not. Direct interaction of UncI protein with c-subunits was suggested from copurification of His-tagged UncI protein and c-subunits, both in the state of c 11-ring and c-monomers. Na+ induced dissociation of His-tagged UncI protein from c 11-ring but not from c-monomers. These results show that UncI is a chaperone-like protein that assists c 11-ring assembly from c-monomers in the membrane.
Footnotes
- ‡To whom correspondence should be addressed at: Chemical Resouces Laboratories, Tokyo Institute of Technology, Nagatsuta 4259, Yokohama 226-8503, Japan. E-mail: myoshida{at}res.titech.ac.jp
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Author contributions: T.S. and M.Y. designed research; T.S., Y.O., N.S., and B.A.F. performed research; T.S. analyzed data; and T.S. and M.Y. wrote the paper.
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The authors declare no conflict of interest.
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This article is a PNAS Direct Submission.
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↵ § In a report of P. modestum F1Fo (12), a band of c 11-ring in SDS/PAGE was observed just below β-subunit, not above α-subunit as observed here. The difference is a result of the gel used. We used gradient gels (10–20% polyacrylamide) in this work. When gels with constant polyacrylamide concentration were used, the band was observed below β-subunit.
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↵ ¶ Plasmid pTRN-ASDS is a derivative of pTR19-ASDS (10). In the plasmid, its lac operator regulation system does not function by unknown mutation, and therefore, the gene introduced is strongly expressed without an inducer, isopropyl-β-thiogalactopyranoside.
- © 2007 by The National Academy of Sciences of the USA
