Abstract
Apocytochromes may represent intermediates in in vivo folding of the holo proteins. Therefore, the question arises, whether apocytochromes have co-operative structure. In the present paper, the holo and apocytochromes b5 and P450cam were studied by differential scanning calorimetry (DSC), limited proteolysis, second derivative spectroscopy, circular dichroism, and size exclusion chromatography.
Apocytochrome b5 is able to undergo a reversible two-state thermal transition. However, transition temperature, denaturational enthalpy and heat capacity change are reduced compared with the holo protein. According to the spectral data, the co-operative structure is mainly based on the core region formed by the residues 1–35 and 79–90. This finding is in full agreement with NMR data (Moore and Lecomte, 1993).
Holo cytochrome P450cam shows in DSC three folding units (domains). Compared with the holo protein, apocytochrome P450cam melting is a two-state process. However, the enthalpy change at thermal melting is reduced from 980 kJ/mol for the holo protein to 135 kJ/mol for the apo form, and the apocytochrome P450cam stability amounts to ΔG = 7.5 kJ/mol.
The apocytochromes are regarded as native-like proteins with extremely low protein stability of about 3 kT, thus showing at the same time some intermediate-like properties. Parts of the molecule produce enhanced partial specific heat capacity and seem not to contribute to the enthalpy and heat capacity change at unfolding. The importance of the properties at in vivo folding are discussed.
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Pfeil, W. (1994). Native-Like Intermediates in Protein Folding. In: Doniach, S. (eds) Statistical Mechanics, Protein Structure, and Protein Substrate Interactions. NATO ASI Series, vol 325. Springer, Boston, MA. https://doi.org/10.1007/978-1-4899-1349-4_4
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DOI: https://doi.org/10.1007/978-1-4899-1349-4_4
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