Side-Chain Interactions in the Plastocyanin−Cytochrome f Complex

Abstract

Cytochrome f and plastocyanin are redox partners in the photosynthetic electron-transfer chain. Electron transfer from cytochrome f to plastocyanin occurs in a specific short-lived complex. To obtain detailed information about the binding interface in this transient complex, the effects of binding on the backbone and side-chain protons of plastocyanin have been analyzed by mapping NMR chemical-shift changes. Cytochrome f was added to plastocyanin up to 0.3 M equiv, and the plastocyanin proton chemical shifts were measured. Out of approximately 500 proton resonances, 86% could be observed with this method. Nineteen percent demonstrate significant chemical-shift changes and these protons are located in the hydrophobic patch (including the copper ligands) and the acidic patches of plastocyanin, demonstrating that both areas are part of the interface in the complex. This is consistent with the recently determined structure of the complex [Ubbink, M., Ejdebäck, M., Karlsson, B. G., and Bendall, D. S. (1998) Structure 6, 323-335]. The largest chemical-shift changes are found around His87 in the hydrophobic patch, which indicates tight contacts and possibly water exclusion from this part of the protein interface. These results support the idea that electron transfer occurs via His87 to the copper in plastocyanin and suggest that the hydrophobic patch determines the specificity of the binding. The chemical-shift changes in the acidic patches are significant but small, suggesting that the acidic groups are involved in electrostatic interactions but remain solvent exposed. The existence of small differences between the present data and those used for the structure may imply that the redox state of the metals in both proteins slightly affects the structure of the complex. The chemical-shift mapping is performed on unlabeled proteins, making it an efficient way to analyze effects of mutations on the structure of the complex.

Structure of the Intermolecular Complex between Plastocyanin and Cytochrome f from Spinach

“…The structures calculated using these pure computational methods suggest the existence of several different orientations used by the two proteins to approach each other. The use of experimental constraints derived from NMR in combination with restrained rigid body molecular mechanics yielded a family of highly similar structures of the transient complex between spinach Pc and turnip Cyt f (22)(23)(24). These structures reveal that van der Waals interactions between the heme region of Cyt f and the hydrophobic patch of Pc bring the two metal ions, iron and copper, within ϳ11 Å of each other.…”

A Brownian dynamics computational study of the interaction of spinach plastocyanin with turnip cytochrome f: the importance of plastocyanin conformational changes

“…See also Crowley and Ubbink (2003) and Ubbink (2004) for a general discussion of the NMR results. These studies document both hydrophobic interactions between the hydrophobic patch surrounding the H87 ligand to the Cu on PC and the corresponding hydrophobic patch surrounding the heme on cyt f. In addition, electrostatic interactions are observed between the negative patch on PC and the positive patch on cyt f (Ejdeback et al 2000). 1 Turnip cyt f. The structure of turnip cyt f is that of Martinez et al (1996) depicted using GRASP (Nicholls and Honig 1991).…”

“…Various lines of evidence including cross-linking (Morand et al 1989), chemical modification of PC (Anderson et al 1987), mutagenesis studies of PC (Lee et al 1995;Kannt et al 1996;Gong et al 2000) and NMR studies of complex formation (Ejdeback et al 2000) implicate electrostatic forces in complex formation between the two proteins. For a more detailed discussion of the evidence for electrostatic interaction between PC and cyt f, see Gross (1996) and Haddadian and Gross (2005).…”

“…NMR has been used to study complex formation between cyt f and various types of higher plant PCs including those from spinach (Ubbink et al 1998;Ejdeback et al 2000), parsley (Crowley et al 2004) and poplar (Lange et al 2005). See also Crowley and Ubbink (2003) and Ubbink (2004) for a general discussion of the NMR results.…”

Electron Transfer and Stability of the Cytochromeb6 f Complex in a Small Domain Deletion Mutant of Cytochrome f

“…However, observations from experiments with in vitro systems yielded different conclusions. The results from the electron transfer experiments carried out in vitro (17,20,21) and in nebulized cells (16) and from molecular dynamic modeling (37)(38)(39) and the solution structure of the complex between cytochrome f and plastocyanin (40,41) provided evidence that the lysine residues from the small domain do interact with plastocyanin. This discrepancy between in vivo and in vitro behaviors remains unresolved.…”