The role of individual lysine residues in the basic patch on turnip cytochrome f for electrostatic interactions with plastocyanin in vitro.
The role of electrostatic interactions in determining the rate of electron transfer between cytochrome f and plastocyanin has been examined in vitro with mutants of turnip cytochrome f and mutants of pea and spinach plastocyanins. Mutation of lysine residues Lys58, Lys65 and Lys187 of cytochrome f to neutral or acidic residues resulted in decreased binding constants and decreased rates of electron transfer to wild-type pea plastocyanin. Interaction of the cytochrome f mutant K187E with the pea plastocyanin mutant D51K gave a further decrease in electron transfer rate, indicating that a complementary charge pair at these positions could not compensate for the decreased overall charge on the proteins. Similar results were obtained with the interaction of the cytochrome f mutant K187E with single, double and triple mutants of residues in the acidic patches of spinach plastocyanin. These results suggest that the lysine residues of the basic patch on cytochrome f are predominantly involved in long-range electrostatic interactions with plastocyanin. However, analysis of the data using thermodynamic cycles provided evidence for the interaction of Lys187 of cytochrome f with Asp51, Asp42 and Glu43 of plastocyanin in the complex, in agreement with a structural model of a cytochrome f-plastocyanin complex determined by NMR.
Gong
X S
XS
Cambridge Centre for Molecular Recognition and Department of Plant Sciences, University of Cambridge, UK.
Wen
J Q
JQ
Fisher
N E
NE
Young
S
S
Howe
C J
CJ
Bendall
D S
DS
Gray
J C
JC
D001224:Aspartic Acid; D001937:Brassica / Q000201:enzymology; D002942:Circular Dichroism; D003580:Cytochromes / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D045348:Cytochromes f; D008239:Lysine; D009682:Magnetic Resonance Spectroscopy; D016297:Mutagenesis, Site-Directed; D010084:Oxidation-Reduction; D010970:Plastocyanin / Q000737:chemistry* / Q000378:metabolism*; D011487:Protein Conformation; D013056:Spectrophotometry, Ultraviolet; D055672:Static Electricity; D013816:Thermodynamics
D003580:Cytochromes; D001224:Aspartic Acid; D010970:Plastocyanin; D045348:Cytochromes f; D008239:Lysine