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

A high thermal stability is observed for the five-stranded alpha-helical co iled-coil domain of cartilage oligomeric matrix protein COMP. It does not u nfold in non-denaturing buffer between 0 and 100 degrees C and thermal dena turation 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 pro file showed deviations from the theor y of Privalov et al. [P.L. Privalov, V. Griko Yu, S. Venyaminov, V.P. Kutys henko, Cold denaturation of myoglobin, J. Mol. Biol. 190 (1986) 487-498] pr obably 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 th e coiled-coil domain of COMP in which glutamine 54 was replaced by isoleuci ne but it still forms pentamer. The melting temperatures in plain buffer fo r the heat denaturation of COMP coiled-coil domain and its mutant obtained by extrapolation to zero molar guanidinium chloride concentration are appro ximately 160 and 220 degrees C, respectively which groups them among the mo st stable proteins. (C) 2000 Elsevier Science B.V. All rights reserved.