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


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. Study holds ProTherm entries: 19063, 19064, 19065, 19066, 19067, 19068, 19069, 19070, 19071, 19072, 19073, 19074, 19075, 19076, 19077, 19078, 19079, 19080, 19081, 19082, 19083, 19084, 19085, 19086, 19087, 19088, 19089, 19090, 19091, 19092, 19093, 19094, 19095, 19096, 19097, 19098, 19099, 19100, 19101, 19102, 19103 Extra Details: circular dichroism; chemical denaturation; fluorescence; NMR; protein stability; thermal denaturation

Submission Details

ID: QremtA4D3

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:51 p.m.

Version: 1

Publication Details
Lidón-Moya MC;Barrera FN;Bueno M;Pérez-Jiménez R;Sancho J;Mateu MG;Neira JL,Protein Sci. (2005) An extensive thermodynamic characterization of the dimerization domain of the HIV-1 capsid protein. PMID:16131662
Additional Information

Number of data points 41
Proteins Gag-Pol polyprotein ; Capsid protein VP24
Unique complexes 1
Assays/Quantities/Protocols Experimental Assay: Tm buffers:phosphate: 25 mM, pH:7.0 ; Experimental Assay: Tm pH:3.0, buffers:acetate: 25 mM ; Experimental Assay: Tm prot_conc:4500 microM, buffers:Sodium acetate: 10 mM, ionic:NaCl: 0.1 M, details:Additives EDTA ( ; Experimental Assay: Tm buffers:Sodium acetate: 10 mM, ionic:NaCl: 0.1 M, prot_conc:2000 microM, details:Additives EDTA ( ; Experimental Assay: Tm buffers:Sodium acetate: 10 mM, prot_conc:1100 microM, ionic:NaCl: 0.1 M, details:Additives EDTA ( ; Experimental Assay: Tm buffers:Sodium acetate: 10 mM, ionic:NaCl: 0.1 M, prot_conc:530 microM, details:Additives EDTA (0 ; Experimental Assay: Tm buffers:Sodium acetate: 10 mM, pH:8.0, ionic:NaCl: 0.1 M, prot_conc:1000 microM, details:Additive ; Experimental Assay: Tm buffers:Sodium acetate: 10 mM, ionic:NaCl: 0.1 M, prot_conc:1000 microM, details:Additives EDTA ( ; Experimental Assay: Tm buffers:Sodium acetate: 10 mM, pH:6.0, ionic:NaCl: 0.1 M, prot_conc:1000 microM, details:Additive ; Experimental Assay: Tm buffers:Unknown: , pH:9.0 ; Experimental Assay: Tm buffers:Unknown: , pH:8.0 ; Experimental Assay: Tm pH:5.0, buffers:Unknown: ; Experimental Assay: Tm buffers:Unknown: , pH:4.0 ; Experimental Assay: Tm buffers:Unknown: , pH:9.0 ; Experimental Assay: Tm pH:6.0, buffers:Unknown: ; Experimental Assay: Tm pH:5.0, buffers:Unknown: ; Experimental Assay: Tm pH:4.0, buffers:Unknown: ; Experimental Assay: Tm prot_conc:20 microM, buffers:Unknown: , pH:9.0 ; Experimental Assay: Tm pH:6.0, prot_conc:20 microM, buffers:Unknown: ; Experimental Assay: Tm prot_conc:20 microM, buffers:Unknown: , pH:5.0 ; Experimental Assay: Tm prot_conc:20 microM, pH:4.0, buffers:Unknown: ; Experimental Assay: Tm buffers:Unknown: , pH:8.0 ; Experimental Assay: Tm buffers:Unknown: , pH:7.0 ; Experimental Assay: Tm prot_conc:20 microM, buffers:Unknown: , pH:8.0 ; Experimental Assay: Tm prot_conc:20 microM, buffers:Unknown: , pH:7.0 ; Experimental Assay: Tm buffers:Unknown: , pH:7.0 ; Experimental Assay: Tm prot_conc:20 microM, buffers:Unknown: , pH:7.0 ; Experimental Assay: Tm buffers:Unknown: , prot_conc:80 microM, pH:7.0, details:Additives dye (100 microM), ; Experimental Assay: Tm prot_conc:20 microM, buffers:Unknown: , pH:9.0, details:Additives dye (100 microM), ; Experimental Assay: Tm prot_conc:20 microM, buffers:Unknown: , pH:8.0, details:Additives dye (100 microM), ; Experimental Assay: Tm prot_conc:20 microM, buffers:Unknown: , pH:7.0, details:Additives dye (100 microM), ; Experimental Assay: Tm pH:6.0, prot_conc:20 microM, buffers:Unknown: , details:Additives dye (100 microM), ; Experimental Assay: Tm buffers:Tris acid: , pH:9.0 ; Experimental Assay: Tm buffers:Tris acid: , pH:8.0 ; Experimental Assay: Tm buffers:Monosodium dihydrogen phosphate: , pH:7.0 ; Experimental Assay: Tm pH:6.0, buffers:Monosodium dihydrogen phosphate: ; Experimental Assay: Tm pH:5.0, buffers:Acetic acid:
Libraries Mutations for sequence MSPTSILDIRQGPKEPFRDYVDRFYKTLRAEQASQEVKNWMTETLLVQNANPDCKTILKALGPGATLEEMMTACQGVGGPGHKARVL
Sequence Assay Result Units