부산대학교 도서관

Ferredoxin-NADP+reductase (FNR) was previously inferred to bind to the cytochrome b6fcomplex in the electron transport chain of oxygenic photosynthesis. In the present study, this inference has been examined through analysis of the thermodynamics of the interaction between FNR and the b6fcomplex. Isothermal titration calorimetry (ITC) was used to characterize the physical interaction of FNR with b6fcomplex derived from two plant sources (Spinacia oleraceaand Zea maize). ITC did not detect a significant interaction of FNR with the b6fcomplex in detergent solution nor with the complex reconstituted in liposomes. A previous inference of a small amplitude but defined FNR-b6finteraction is explained by FNR interaction with micelles of the undecyl β-D maltoside (UDM) detergent micelles used to purify b6f. Circular dichroism, employed to analyze the effect of detergent on the FNR structure, did not reveal significant changes in secondary or tertiary structures of FNR domains in the presence of UDM detergent. However, thermodynamic analysis implied a significant decrease in an interaction between the N-terminal FAD-binding and C-terminal NADP+-binding domains of FNR caused by detergent. The enthalpy, ΔHo, and the entropy, ΔSo, associated with FNR unfolding decreased four-fold in the presence of 1 mM UDM at pH 6.5. In addition to the conclusion regarding the absence of a binding interaction of significant amplitude between FNR and the b6fcomplex, these studies provide a precedent for consideration of significant background protein-detergent interactions in ITC analyses involving integral membrane proteins.