Effects of heme on the thermal stability of mesophilic and thermophilic cytochromes c: Comparison between experimental and theoretical results

Abstract

We have recently proposed a measure of the thermal stability of a protein: the water-entropy gain at 25 °C upon folding normalized by the number of residues, which is calculated using a hybrid of the angle-dependent integral equation theory combined with the multipolar water model and the morphometric approach. A protein with a larger value of the measure is thermally more stable. Here we extend the study to analyses on the effects of heme on the thermal stability of four cytochromes c (PA c(551), PH c(552), HT c(552), and AA c(555)) whose denaturation temperatures are considerably different from one another despite that they share significantly high sequence homology and similar three-dimensional folds. The major conclusions are as follows. For all the four cytochromes c, the thermal stability is largely enhanced by the heme binding in terms of the water entropy. For the holo states, the measure is the largest for AA c(555). However, AA c(555) has the lowest packing efficiency of heme and the apo polypeptide with hololike structure, which is unfavorable for the water entropy. The highest stability of AA c(555) is ascribed primarily to the highest efficiency of side-chain packing of the apo polypeptide itself. We argue for all the four cytochromes c that due to covalent heme linkages, the number of accessible conformations of the denatured state is decreased by the steric hindrance of heme, and the conformational-entropy loss upon folding becomes smaller, leading to an enhancement of the thermal stability. As for the apo state modeled as the native structure whose heme is removed, AA c(555) has a much larger value of the measure than the other three. Overall, the theoretical results are quite consistent with the experimental observations (e.g., at 25 °C the α-helix content of the apo state of AA c(555) is almost equal to that of the holo state while almost all helices are collapsed in the apo states of PA c(551), PH c(552), and HT c(552)).

Essential roles of protein-solvent many-body correlation in solvent-entropy effect on protein folding and denaturation: Comparison between hard-sphere solvent and water

“…64 Protein folding is accompanied by a solvent-entropy gain ∆S (∆S > 0) and a loss of the protein conformational entropy ∆S C (∆S C < 0). Above 298 K, ∆S and |∆S C | are decreasing and increasing functions of T, respectively.…”

Extracellular Electron Transfer to Fe(III) Oxides by the Hyperthermophilic Archaeon Geoglobus ahangari via a Direct Contact Mechanism

“…Most of the redox-active proteins catalyzing this last step of electron transfer in mesophilic Fe(III) reducers are heme-containing c cytochromes (9). These proteins are especially suited for electron transfer at high temperatures because the covalent attachment of heme groups to proteins stabilizes their secondary structure and increases their thermal stability (42). However, dithionite-reduced versus air-oxidized spectral analyses of resting cell suspensions of G. ahangari failed to identify the spectral signatures characteristic of c-type cytochromes (25).…”

Morphometric approach to thermodynamic quantities of solvation of complex molecules: Extension to multicomponent solvent

“…We have been quite successful in elucidating pressure, 14,15,39,40 thermal, 16,17 and cold 8,9 denaturation with the emphasis on the entropic components of the system free energy. Our principal concern in the next stage is the elucidation of protein denaturation caused by the cosolvent (e.g., urea) addition on the basis of the entropic components.…”

“…The MA or similar approaches have been applied to a variety of problems where complexly shaped, solvophobic solutes are treated, [10][11][12][13] or where the solvophobic effect plays important roles. In particular, we have been quite successful in elucidating folding/unfolding mechanisms of proteins, [5][6][7][8][9][14][15][16][17] discriminating the native fold of a protein from a number of misfolded decoys, [18][19][20] and uncovering the rotation mechanism of a motor protein. 21 In these problems, the solvation entropy is the key quantity calculated by modeling a protein with a fixed structure as a set of fused hard spheres.…”

“…In studies on the unfolding mechanisms, the SFE or related quantities of solvation thermodynamics must be calculated for a number of different structures of a protein, and the use of an efficient method, such as the MA, is highly necessitated. We have developed physical pictures of pressure, 14,15 cold, 8,9 and thermal 16,17 denaturating by employing the MA. The remaining problem is the denaturation caused by the addition of cosolvents such as urea.…”