The Alkali Molten Globule State of Horse Ferricytochrome c: Observation of Cold Denaturation

Analysis of the pH-dependent stability and millisecond folding kinetics of horse cytochrome c

“…S3 H) fails the basic test for accumulation of equilibrium structural intermediate to a detectable level. A few earlier spectroscopic and calorimetric studies have revealed that at low ionic strength, the ratio between van't Hoff and calorimetric enthalpy for thermal denaturation of Ferricyt c under conditions of low pH (pH 2.2e3.6), neutral pH (pH~7.0) and high pH (pH 12.5e13.0) is close to one [87,100,101,103], indicating that the thermal denaturation of Ferricyt c under these conditions of pH is two state [69,93,95,98]. An another previous spectroscopic and calorimetric study has shown that at pH 7.0, the ratio between van't Hoff and calorimetric enthalpy for thermal denaturation of Ferrocyt c is close to one [99], indicating that the thermal denaturation of Ferrocyt c at neutral pH is also two state.…”

“…The decrease in m g -values at alkaline pH relative to neutral pH is likely due to (i) relatively small solvent accessible area in the denatured state of protein at pH ! 11.5, (ii) deviation from two-state folding [95], and (iii) denaturant unfolded cyt c is more compact at alkaline pH than at neutral pH [87,96]. The fluorescence emission intensity of denaturant unfolded cyt c at pH 13 is at least 2e5 fold less relative to pH 7 [87,96], which is possibly due to either quenching by hydroxyl ions or ionization of the phenolic hydroxyl group of tyrosine at pH 13.…”

“…It is likely that the partial interactions of cyt c molecules occur at neutral pH and that may play an important role for not showing the high reversibility in thermal denaturations of Ferricyt c and Ferrocyt c at pH 7.0. The observed irreversibility in thermal denaturation of cyt c may depend on pH and which can affect the thermodynamic parameters associated with the pHdependent thermal denaturation of Ferricyt c and Ferrocyt c. A few earlier spectroscopic and calorimetric studies have revealed that at low ionic strength, the thermal denaturation of Ferricyt c at low pH (2.2e3.6) and high pH (12.8e13.0) is reversible and two state [87,100,101]. At low pH (2.2e3.6), a high number of positive charges may prevent the intermolecular interactions of cyt c molecules [102] and that may play a key role for the reversible twostate behavior in the thermal denaturation of Ferricyt c at low pH (2.2e3.6) [102].…”

A fully atomistic computer simulation study of cold denaturation of a β-hairpin

“…As the weakening hydrophobic force with decreasing temperature appears to be a key driving force governing cold denaturation, investigations of cold denaturation using either experimental or computational methods can provide new insights into the nature of hydrophobic effects. The thermodynamics of cold denaturation has been well-characterized experimentally for several globular proteins by calorimetric 3 , circular dichroism 4,5 , and nuclear magnetic resonance (NMR) 6,7 experiments. Notably, the colddenatured structure of ubiquitin was fully determined using a pressure-assisted NMR method 8 .…”

Distribution and Transition of Native and Completely Unfolded Conformations in the Unfolding of Bovine Heart Cytochrome c Induced by Urea and Guanidine Hydrochloride

“…Wherein, horse heart cytochrome c may be the most used cytochrome c molecules. For example, Harrington studied the unfolding of horse ferricytochrome c in the presence of several inorganic salts under a variety of denaturing conditions by means of viscosity measurements and absorbance changes in the Soret and visible regions; 5 Nieman et al studied the 13 C NMR spectroscopy of acetyltyrosyl-guanidinated horse heart cytochrome c; 6 Hagihara et al compared the stability of the native and acidic molten globule states of horse ferricytochrome c against heat, urea and guanidine hydrochloride using the intact species and species modified by various degrees of acetylation of the lysyl ε-amino groups; 7 Zhu et al investigated the conformational transition of horse heart cytochrome c induced by bromopyrogal red in very low concentration by dynamic spectro-electrochemical technique; 8 Zhang et al studied the denaturation of horse heart cytochrome c induced by bromopyrogal red by scanning tunnelling microscopy on the electrochemically pretreated highly oriented pyrolytic graphite surface; 9 Yang et al studied conformational changes of horse heart cytochrome c determined by matrix-assisted laser desorption mass spectrometry; 10 Moosavi-Movahedi et al studied the differential scanning calorimetry of the molten globule state of horse heart cytochrome c induced by sodium n-dodecyl sulfate; 11 Krylov et al studied the denaturation and renaturation of horse heart cytochrome c immobilized on gold electrodes in DMSO-containing buffers; 12 Kumar et al studied the alkali molten globule state of horse ferricytochrome c in "cold denaturation"; 13 Latypov et al studied the equilibrium and kinetic properties of reduced horse heart cytochrome c in the presence of carbon monoxide; 14 Baker and Heller studied the characteristics of equine heart cytochrome c in aqueous solutions of the fully water-miscible IL 1-butyl-3-methyl-imidazolium chloride by small-angle neutron and X-ray scattering; 15 Liu and Konermann characterized the solution-phase properties of horse heart cytochrome c after heat exposure by electrospray ionization mass spectrometry in conjunction with hydrogen/deuterium exchange and optical spectroscopy. 16 Like horse heart cytochrome c, cytochrome c from bovine heart is also a globular protein molecule.…”