Factors that determine the unusually low reduction potential of cytochrome c550 in cyanobacterial photosystem II

John S. Vrettos, Michael J. Reifler, Olaf Kievit, K. V. Lakshmi, Julio C. De Paula, Gary W. Brudvig

Research output: Contribution to journalArticlepeer-review

27 Citations (Scopus)


A new purification protocol for cytochrome c550 (cyt c550]) from His-tagged Synechocystis PCC 6803 photosystem II (PSII) was developed which allows the protein to be isolated in high yield and purity. Electron paramagnetic resonance spectroscopy of cyt c550, both free in solution and in intact PSII preparations, yields identical spectra with g values at 1.50, 2.23, and 2.87, which are characteristic for a ferric low-spin bis-histidine coordinated heme. The resonance Raman spectrum of the isolated protein exhibits features characteristic of bis-histidine axial ligation of the iron and a slight ruffling of the heme macrocycle. Together, these results indicate that the heme structure is not very different from most c-type cytochromes, and thus the structure of the heme does not account for its unusually low reduction potential. A direct electrochemical measurement of the reduction potential was performed using square wave voltammetry on a pyrolytic graphite edge electrode, yielding E1/2 = -108 mV (vs. NHE) with a peak separation of 5 mV. This value is 150 mV more positive than that previously measured by redox titrations. Because the behavior of the protein in the electrochemistry experiments is indicative of adsorption to the electrode surface, we surmise that binding of the protein to the electrode excludes solvent water from the heme-binding site. We conclude that the degree of solvent exposure makes a significant contribution to the heme reduction potential. Similarly, the binding of cyt c550 to PSII may also reduce the solvent exposure of the heme, and so the direct electrochemical value of the reduction potential may be relevant to the protein in its native state.

Original languageEnglish
Pages (from-to)708-716
Number of pages9
JournalJournal of Biological Inorganic Chemistry
Issue number7
Publication statusPublished - 2001


  • Cytochrome c
  • EPR spectroscopy
  • Photosystem II
  • Protein electrochemistry
  • Resonance Raman spectroscopy

ASJC Scopus subject areas

  • Biochemistry
  • Inorganic Chemistry

Fingerprint Dive into the research topics of 'Factors that determine the unusually low reduction potential of cytochrome c<sub>550</sub> in cyanobacterial photosystem II'. Together they form a unique fingerprint.

Cite this