Conformation and stability of recombinant HIV-1 capsid protein p24 (rp24).

Conformation and stability of the recombinant protein HIV-1 rp24 were analyzed by circular dichroism, fluorescence spectroscopy and differential scanning calorimetry under different solvent conditions. From circular dichroism measurements, HIV-1 rp24 at pH 5.8 can be classified as an all alpha-helical protein. A fluorescence maximum of about 330 nm indicates a predominantly hydrophobic environment of the five tryptophan residues. The GdnHCl-induced unfolding curves monitored by CD and fluorescence are sigmoidal and single phasic and the midpoints of transitions are independent on the protein concentration. For the calculation of free energy of unfolding delta GuH2O a 'two-state' model was applied. The calculated values are between 18 and 24 kJ/mol and thus on the lower limit of the conformational stability of globular proteins. Melting experiments at pH 5.8 are impaired by a strong irreversible aggregation at higher temperatures. However, at pH 3.0 and in the presence of 0.1% (w/v) ocytl beta-glucopyranoside the melting curves show a large degree of reversibility with a Tm value of 38 degrees C and a molar enthalpy change delta Hm of 218 kJ/mol. At pH < 2.5 HIV-1 rp24 can adopt a new conformation which is characterized by a high alpha-helical content, a strongly decreased CD in the aromatic region, a red-shift of the fluorescence spectrum and a strong binding of ANS. These spectral features of the acid-induced conformational state are similar to those obtained for molten globule-like folding states. HIV-1 rp24 unfolds cooperatively at pH 2.0 in the concentration range of about 1.5-3.0 M GdnHCl. The calculated values delta GuH2O at pH 2.0 of about 12 kJ/mol are significantly decreased in comparison to the delta GuH2O values of the protein at pH 5.8.