title: Non-two-state thermal denaturation of ferricytochrome c at neutral and slightly acidic pH values.
abstract: Thermal denaturation of ferricytochrome c (cyt c) has been methodically studied by absorbance, fluorescence, circular dichroism spectroscopy, viscosimetry and differential scanning calorimetry in pH range from pH 3.5 to 7.5. Thermal transitions have been monitored by intrinsic local probes of heme region such as absorbance at Soret, 620nm and 695nm bands and circular dichroism signals at 417nm. Global conformational changes were analyzed by circular dichroism signal at 222nm, fluorescence of the single tryptophan, reduced viscosity and differential scanning calorimetry. We show that cyt c thermal denaturation above pH ~5 can be described by an apparent two-step transition in which the heme iron stays in a low-spin state. The thermal denaturations of cyt c below pH ~5 proceed in one step to an unfolded highly compact form with a high-spin state of the heme iron. Cyt c conformational plasticity is discussed in regard to its physiological functions.
authors.affiliation: Department of Biochemistry, P.J. Šafárik University, Moyzesova 11, 040 01 Košice, Slovakia.
authors.forename: Rastislav
authors.initials: R
authors.lastname: Varhač
authors.affiliation: Department of Biophysics, Institute of Experimental Physics Slovak Academy of Sciences, Watsonova 47, 040 01 Košice, Slovakia.
authors.forename: Dagmar
authors.initials: D
authors.lastname: Sedláková
authors.affiliation: Department of Medical and Clinical Biochemistry, Faculty of Medicine, P.J. Šafárik University, Trieda SNP 1, 040 11 Košice, Slovakia.
authors.forename: Marek
authors.initials: M
authors.lastname: Stupák
authors.affiliation: Department of Biochemistry, P.J. Šafárik University, Moyzesova 11, 040 01 Košice, Slovakia; Centre for Interdisciplinary Biosciences, P.J. Šafárik University, Jesenná 5, 040 01 Košice, Slovakia. Electronic address: erik.sedlak@upjs.sk.
authors.forename: Erik
authors.initials: E
authors.lastname: Sedlák
mesh_terms: D002152:Calorimetry, Differential Scanning; D002942:Circular Dichroism; D045304:Cytochromes c / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D011487:Protein Conformation; D011489:Protein Denaturation; D013696:Temperature; D014783:Viscosity
chemical_list: D045304:Cytochromes c
keywords: Alkaline state; Conformational change; Ligand exchange; Protein stability; Thermal denaturation