The unusually stable coiled-coil domain of COMP exhibits cold and heat denaturation in 4–6 M guanidinium chloride

Abstract

A high thermal stability is observed for the five-stranded α-helical coiled-coil domain of cartilage oligomeric matrix protein COMP. It does not unfold in non-denaturing buffer between 0 and 100°C and thermal denaturation is only achieved at high concentrations of guanidinium chloride (4–6 M). In these solutions the protein structure is lost at decreasing (cold denaturation) and increasing temperatures (heat denaturation). In the cold denaturation region, the melting profile showed deviations from the theory of Privalov et al. [P.L. Privalov, V. Griko Yu, S. Venyaminov, V.P. Kutyshenko, Cold denaturation of myoglobin, J. Mol. Biol. 190 (1986) 487–498] probably due to deviations from a two-state mechanism. High thermal stability as well as cold and heat denaturation was also observed for a mutant of the coiled-coil domain of COMP in which glutamine 54 was replaced by isoleucine but it still forms pentamer. The melting temperatures in plain buffer for the heat denaturation of COMP coiled-coil domain and its mutant obtained by extrapolation to zero molar guanidinium chloride concentration are approximately 160 and 220°C, respectively, which groups them among the most stable proteins.