Stern-volmer in reverse: 2:1 Stoichiometry of the cytochrome c-cytochrome c peroxidase electron-transfer complex

Jian S. Zhou, Brian M. Hoffman*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

85 Scopus citations

Abstract

A reverse protocol for measurements of molecular binding and reactivity by excited-state quenching has been developed in which the quencher, held at a fixed concentration, is titrated by a photoexcitable probe molecule whose decay is monitored. The binding stoichiometries, affinities, and reactivities of the electron-transfer complexes between cytochrome c (Cc) and cytochrome c peroxidase (CcP) were determined over a wide range of ionic strengths (4.5 to 118 millimolar) by the study of photoinduced electron-transfer quenching of the triplet excited state of zinc-substituted Cc (ZnCc) by Fe3+CcP. The 2:1 stoichiometry seen for the binding of Cc to CcP at low ionic strength persists at the physiologically relevant ionic strengths and likely has functional significance. Analysis of the stoichiometric binding and rate constants confirms that one surface domain of CcP binds Cc with a high affinity but with poor electron-transfer quenching of triplet-state ZnCc, whereas a second binds weakly but with a high rate of electron-transfer quenching.

Original languageEnglish (US)
Pages (from-to)1693-1696
Number of pages4
JournalScience
Volume265
Issue number5179
DOIs
StatePublished - 1994

ASJC Scopus subject areas

  • General

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