Hybrid Fo complexes of the ATP synthases of spinach chloroplasts and Escherichia coli. Immunoprecipitation and mutant analyses.
Hybrid Fo complexes of the ATP synthases of spinach chloroplast (CFo) and Escherichia coli (EFo) were investigated. Immunoprecipitations with polyclonal antibodies against the different Fo subunits clearly revealed that hybrid Fo complexes derived from CFo subunit III and EFo subunits a and b were formed in vivo. In addition, the ATPase activities of the hybrid ATP synthase, measured in everted cytoplasmic membranes of an atpE mutant strain transformed with the atpH gene coding for CFo III, were comparable to activities obtained for the same mutant strain complemented with the atpE gene (EFo c). Nevertheless, CFo III was not able to replace EFo c functionally, since the strain containing the hybrid ATP synthase was not able to grow on succinate. In order to investigate the reason for this lack of function, hybrid proteolipids of CFo III and EFo c were constructed. Only a chimaeric protein comprising the seven N-terminal amino acid residues from CFo III and the remaining part of EFo c was able to replace wild-type EFo c, whereas hybrid proteins with 13 and 33 N-terminal amino acids of CFo III were not functional. The results suggested that a network of interactions between the subunits essential for proton translocation and/or coupling of the F1 part exists, which was optimized for each species during evolution, although the overall structure of FoF1 complexes has been conserved.
Burkovski
A
A
Universität Osnabrück, Fachbereich Biologie/Chemie, Germany.
Deckers-Hebestreit
G
G
Altendorf
K
K
D000595:Amino Acid Sequence; D002736:Chloroplasts / Q000201:enzymology; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics*; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D016133:Polymerase Chain Reaction; D011233:Precipitin Tests; D011510:Proteolipids / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D006180:Proton-Translocating ATPases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D018724:Spinacia oleracea / Q000201:enzymology* / Q000235:genetics*
D011510:Proteolipids; D011993:Recombinant Fusion Proteins; D006180:Proton-Translocating ATPases