Identification of the P-loop lysine as a metal ligand in the absence of nucleotide at catalytic site 3 of chloroplast F1-ATPase from Chlamydomonas reinhardtii.
Site-directed mutations were made to the phosphate-binding loop lysine in the beta-subunit of the chloroplast F(1)-ATPase in Chlamydomonas reinhardtii (betaK167) to investigate the participation of this residue in the binding of metal to catalytic site 3 in the absence of nucleotide. The cw-EPR spectra of VO(2+) bound to site 3 of CF(1)-ATPase from wild type and mutants revealed changes in metal ligation resulting from mutations to betaK167. The three-pulse ESEEM spectrum of the wild-type CF(1)-ATPase with VO(2+) bound to site 3 shows an equatorially coordinating (14)N from an amine. The ESEEM spectra of the mutants do not show evidence of an equatorially coordinating amine group. The results presented here show that, in the absence of nucleotide, betaK167 is a ligand to the metal bound at catalytic site 3, suggesting a regulatory role for the P-loop lysine in addition to its known role in catalysis.
Crampton
D J
DJ
The Center for the Study of Early Events in Photosynthesis, Arizona State University, Tempe 85287-1601, USA.
LoBrutto
R
R
Frasch
W D
WD
D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D020134:Catalytic Domain / Q000235:genetics; D016825:Chlamydomonas reinhardtii / Q000201:enzymology* / Q000235:genetics; D002736:Chloroplasts / Q000201:enzymology / Q000235:genetics; D004578:Electron Spin Resonance Spectroscopy; D008024:Ligands; D008239:Lysine / Q000235:genetics / Q000378:metabolism*; D008274:Magnesium / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D010100:Oxygen / Q000378:metabolism; D006180:Proton-Translocating ATPases / Q000235:genetics / Q000378:metabolism*; D014639:Vanadium / Q000378:metabolism*
D008024:Ligands; D014639:Vanadium; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D006180:Proton-Translocating ATPases; D008274:Magnesium; D008239:Lysine; D010100:Oxygen