Fluorescence resonance energy transfer mapping of subunit delta in spinach chloroplast F1 ATPase.
Despite the considerable progress in the field of F0F1-ATPases caused by solving the 2.8-A structure of mitochondrial F1 ATPase [Abrahams, J. P., Leslie, A. G. W., Lutter, R. & Walker, J. E. (1994) Nature 370, 621-628], little is known about the position and function of the enzyme's small subunits which were not resolved in the X-ray analysis. We have previously genetically engineered Cys residues into the delta subunit of chloroplast F1 and used these mutant subunits in cross-linking studies [Lill, H., Hensel, F., Junge, W. & Engelbrecht, S. (1996) J. Biol. Chem. 271, 32737-32742]. In this work, various fluorophores have been introduced into the mutant delta subunits and used in fluorescence-resonance energy-transfer measurements. The resulting distances were fitted into the framework of existing data. Subunit delta was found to be located between two alpha/beta couples, stretching from the level of the nucleotide binding sites up to a position close to the N-termini of subunits alpha and beta. These results corroborate and further refine the previously found location of spinach CF1 delta at the periphery and membrane-distal part of CF1, where it may constitute a part of a stator in the rotatory machinery of F0F1.
Engelbrecht
S
S
Universität Osnabrück, Abteilung Biophysik, Germany.
Giakas
E
E
Marx
O
O
Lill
H
H
D015081:2-Naphthylamine / Q000031:analogs & derivatives / Q000378:metabolism; D001665:Binding Sites / Q000502:physiology; D002736:Chloroplasts / Q000737:chemistry*; D003374:Coumarins / Q000378:metabolism; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005456:Fluorescent Dyes / Q000378:metabolism; D008958:Models, Molecular; D016296:Mutagenesis / Q000235:genetics; D010940:Plant Proteins / Q000737:chemistry; D011487:Protein Conformation; D006180:Proton-Translocating ATPases / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D018724:Spinacia oleracea / Q000201:enzymology*
D003374:Coumarins; D005456:Fluorescent Dyes; D010940:Plant Proteins; D011994:Recombinant Proteins; C038518:acrylodan; D015081:2-Naphthylamine; D006180:Proton-Translocating ATPases; D003545:Cysteine