Intergenic suppression in a beta subunit mutant with defective assembly in Escherichia coli F1ATPase. Second-site mutation in the alpha subunit.
Substitution of Leu-40 by Pro in the beta subunit (beta L40P) of Escherichia coli F1-ATPase caused a decrease in the amount of the alpha and beta subunits on the membranes. A revertant strain, Re50, carrying no suppression mutations in the uncD gene encoding the beta subunit, was isolated from the beta L40P mutant. The uncA gene from this revertant was amplified by PCR, and cloned into an expression plasmid. The expression plasmid carrying the uncA gene from the revertant was used for genetic suppression assays. The suppression mutation in Re50 was in the alpha subunit, and it recovered the assembly of the alpha and beta subunits into the F1F0 complex and the ATPase activity to 50% that of the wild type. In Re50, Leu-111 was substituted by Gln in the alpha subunit. These results suggest that the regions including Leu-40 in the beta subunit and Leu-111 in the alpha subunit are located close together and interact with each other, either directly or indirectly.
Miki
J
J
Department of Biotechnology, Faculty of Engineering Sciences, Okayama University, Japan.
Tsugumi
S
S
Ikeda
H
H
Kanazawa
H
H
D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids; D016133:Polymerase Chain Reaction; D006180:Proton-Translocating ATPases / Q000737:chemistry / Q000235:genetics*; D015183:Restriction Mapping; D013489:Suppression, Genetic
D046911:Macromolecular Substances; D006180:Proton-Translocating ATPases