Use of chemical modification to study the relationship between activity and net protein charge of the photosystem I...

The net charge on the photosystem I core complex of Shiozawa et al. (Shiozawa, J. A., Alberte, R. S., & Thornber, J. P. (1974) Arch. Biochem. Biophys. 165, 388-397) has been altered by using a water-soluble carbodiimide to form an amide bond between protein carboxyl groups and an amino group of ethylenediamine. This process replaces negatively charged carboxyl groups with positively charged free amino groups. Six hundred moles of ethylenediamine was incorporated per mole of P700 reaction center. The modification of the complex shifted the isoelectric pH of photosystem I from 5.0 to 9.5 without affecting the total amount of P700. The modified complex exhibited an increase in energy transfer from light-harvesting chlorophyll alpha molecules to the reaction center. This phenomenon has been previously observed upon addition of Mg2+ ions to the complex (Gross, E. L., & Grenier, J. (1978) Arch. Biochem. Biophys. 187, 387-398). The modification replaces the divalent cation requirement for electron donation to P700 by plastocyanin and lowered the Km for plastocyanin binding to 2.0 microM, compared to 32 microM for control photosystem I in the presence of Mg2+. In addition, the modification lowered the Km for the negatively charged electron donors dichlorophenolindophenol and ascorbate. These results suggest that changing the charge on the photosystem I complex from negative to positive stimulates both light utilization and electron transfer from electron donors to P700. We suggest that cation regulation of photosystem I activity occurs by a process in which cations alter the charge of the local environment around the complex.