An extensive thermodynamic characterization of the dimerization domain of the HIV-1 capsid protein

Protein Sci. 2005 Sep;14(9):2387-404. doi: 10.1110/ps.041324305.

Abstract

The type 1 human immunodeficiency virus presents a conical capsid formed by several hundred units of the capsid protein, CA. Homodimerization of CA occurs via its C-terminal domain, CA-C. This self-association process, which is thought to be pH-dependent, seems to constitute a key step in virus assembly. CA-C isolated in solution is able to dimerize. An extensive thermodynamic characterization of the dimeric and monomeric species of CA-C at different pHs has been carried out by using fluorescence, circular dichroism (CD), absorbance, nuclear magnetic resonance (NMR), Fourier transform infrared (FTIR), and size-exclusion chromatography (SEC). Thermal and chemical denaturation allowed the determination of the thermodynamic parameters describing the unfolding of both CA-C species. Three reversible thermal transitions were observed, depending on the technique employed. The first one was protein concentration-dependent; it was observed by FTIR and NMR, and consisted of a broad transition occurring between 290 and 315 K; this transition involves dimer dissociation. The second transition (Tm approximately 325 K) was observed by ANS-binding experiments, fluorescence anisotropy, and near-UV CD; it involves partial unfolding of the monomeric species. Finally, absorbance, far-UV CD, and NMR revealed a third transition occurring at Tm approximately 333 K, which involves global unfolding of the monomeric species. Thus, dimer dissociation and monomer unfolding were not coupled. At low pH, CA-C underwent a conformational transition, leading to a species displaying ANS binding, a low CD signal, a red-shifted fluorescence spectrum, and a change in compactness. These features are characteristic of molten globule-like conformations, and they resemble the properties of the second species observed in thermal unfolding.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anilino Naphthalenesulfonates / chemistry
  • Anilino Naphthalenesulfonates / metabolism
  • Anisotropy
  • Capsid Proteins / chemistry*
  • Capsid Proteins / metabolism
  • Chromatography, Liquid / methods
  • Circular Dichroism
  • Dimerization
  • Fluorescence
  • HIV-1 / chemistry*
  • Hydrogen-Ion Concentration
  • Magnetic Resonance Spectroscopy
  • Protein Conformation
  • Protein Denaturation
  • Protein Folding
  • Protein Structure, Tertiary
  • Spectroscopy, Fourier Transform Infrared
  • Thermodynamics*
  • Tryptophan / chemistry
  • Tyrosine / chemistry

Substances

  • Anilino Naphthalenesulfonates
  • Capsid Proteins
  • Tyrosine
  • 1-anilino-8-naphthalenesulfonate
  • Tryptophan