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Published February 9, 1999 | public
Journal Article

Role of Ligand Substitution in Ferrocytochrome c Folding

Abstract

The ligand substitutions that occur during the folding of ferrocytochrome c [Fe(II)cyt c] have been monitored by transient absorption spectroscopy. The folding reaction was triggered by photoinduced electron transfer to unfolded Fe(III)cyt c in guanidine hydrochloride (GuHCl) solutions. Assignments of ligation states were made by reference to the spectra of the imidazole and methionine adducts of N-acetylated microperoxidase 8. At pH 7, the heme in unfolded Fe(II)cyt c is ligated by native His18 and HisX (X = 26, 33) residues. The native Met80 ligand displaces HisX only in the last stages of folding. The ferroheme is predominantly five-coordinate in acidic solution; it remains five-coordinate until the native methionine binds the heme to give the folded protein (the rate of the methionine binding step is 16 ± 5 s^(-1) at pH 5, 3.2 M GuHCl). The evidence suggests that the substitution of histidine by methionine is strongly coupled to backbone folding.

Additional Information

© 1999 American Chemical Society. Received August 11, 1998; Revised Manuscript Received October 21, 1998. Supported by the National Science Foundation (MCB 9630465) and the National Institutes of Health (GM18660 to J.R.T.). We thank Gary Mines and Torbjörn Pascher for assistance with certain experiments and for helpful discussions.

Additional details

Created:
August 19, 2023
Modified:
October 18, 2023