Structure-Function of the Cytochrome b6f Complex in Oxygenic Photosynthesis: Molecular Control of Electron Transport and Thermodynamic Analysis of the Interaction of a Proposed Protein Ligand
2020-07-31T18:15:45Z (GMT) by
In the first study presented here, the 2.5 Å crystal structure1 of the cytochrome b6f complex obtained from the cyanobacterium Nostoc sp. PCC 7120 (pdb 4OGQ) was used as a guide for modification by site-directed mutagenesis in the cyanobacterium Synechococcus sp. PCC 7002 of the rate-limiting step in the central electron transport/proton translocation chain of oxygenic photosynthesis. This step is associated with the oxidation and deprotonation of plastoquinol on the electrochemically positive (p) side of the membrane. The mutagenesis strategy is based on structure studies of the b6f complex in the absence and presence of quinol analogue inhibitors which bind and inhibit electron transport on the p-side of the thylakoid membrane. The strategy focused on two conserved prolines located on the p-side of the F-helix, proximal to the C-helix, in subunit IV of the seven subunit cytochrome b6f complex. These prolines, residues 105 and 112 in the F-helix, are seen in the crystal structure to cause a bend in this helix away from the C-helix in the cytochrome b subunit. Thus, they are predicted to increase the portal aperture for the plastoquinol generated in the photosystem II reaction center complex that serves as the electron-proton donor to the [2Fe-2S] iron-sulfur protein and the pside b-heme. Changing the two prolines to alanine resulted in a decrease of 30-50 % in the logphase growth rate of the cell culture and reduction of photo-oxidized cytochrome f. The second study examines the binding thermodynamics of the cytochrome b6f complex and a purposed binding partner, PGRL1, using isothermal titration calorimetry. Proton Gradient Regulation-Like 1 (PGRL1) is thought to be necessary for efficient cyclic electron transfer, however, it’s mechanistic role is unknown. Here we examined for PGRL1 and cytochrome b6f complex binding and found there was no detectable interaction, indicating that PGRL1 is not a direct quinone/cyt b6f electron cofactor.