Alan E. Senior1,2,3,4, J. Allan Downie1,2,3,4, G B Cox1,2,3,4, F. Gibson1,2,3,4, L Langman1,2,3,4, D R Fayle1,2,3,4
1Department of Biochemistry
2Department of Biochemistry, John Curtin School of Medical Research, Australian National University, Canberra, A.C.T. 2601, Australia
3Department of Biochemistry, Scripps Clinic and Research Foundation, La Jolla, CA 92307, U.S.A.
4School of Medicine and Dentistry, University of Rochester, 601 Elmwood Avenue, Rochester, NY 14642, U.S.A.
Tóm tắt
Four mutant strains of Escherichia coli which lack membrane-bound adenosine triphosphatase activity were shown by genetic-complementation tests to carry mutations in the uncA gene. A soluble inactive F1-ATPase aggregate was released from the membranes of three of the uncA mutant strains by low-ionic-strength washing, and purified by procedures developed for the purification of F1-ATPase from normal strains. Analysis of the subunit structure by two-dimensional gel electrophoresis indicated that the F1-ATPase in strains carrying the uncA401 or uncA453 alleles had a subunit structure indistinguishable from normal F1-ATPase. In contrast, the F1-ATPase from the strain carrying the uncA447 allele contained an alpha-subunit of normal molecular weight, but abnormal net charge. Membranes from strains carrying the uncA450 allele did not have F1-ATPase aggregates that could be solubilized by low-ionic-strength washing. However, a partial dipolid strain carrying both the uncA+ and uncA450 alleles formed an active F1-ATPase aggregate which could be solubilized by low-ionic-strength washing of the membranes and which contained two types of alpha-subunit, one of which was normal and the other had abnormal net charge. It is concluded that the uncA gene codes for the alpha-subunit of the adenosine triphosphatase.
