UV and X-ray structural studies of a 101-residue long Tat protein from a HIV-1 primary isolate and of its mutated, detoxified, vaccine candidate.
The 101-residue long Tat protein of primary isolate 133 of the human immunodeficiency virus type 1 (HIV-1), wt-Tat(133) displays a high transactivation activity in vitro, whereas the mutant thereof, STLA-Tat(133), a vaccine candidate for HIV-1, has none. These two proteins were chemically synthesized and their biological activity was validated. Their structural properties were characterized using circular dichroism (CD), fluorescence emission, gel filtration, dynamic light scattering, and small angle X-ray scattering (SAXS) techniques. SAXS studies revealed that both proteins were extended and belong to the family of intrinsically unstructured proteins. CD measurements showed that wt-Tat(133) or STLA-Tat(133) underwent limited structural rearrangements when complexed with specific fragments of antibodies. Crystallization trials have been performed on the two forms, assuming that the Tat(133) proteins might have a better propensity to fold in supersaturated conditions, and small crystals have been obtained. These results suggest that biologically active Tat protein is natively unfolded and requires only a limited gain of structure for its function.
Foucault
Marine
M
IBCP UMR CNRS Université de Lyon, IFR, France.
Mayol
Katia
K
Receveur-Bréchot
Véronique
V
Bussat
Marie-Claire
MC
Klinguer-Hamour
Christine
C
Verrier
Bernard
B
Beck
Alain
A
Haser
Richard
R
Gouet
Patrice
P
Guillon
Christophe
C
D016915:AIDS Vaccines / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D000595:Amino Acid Sequence; D002850:Chromatography, Gel; D002942:Circular Dichroism; D018360:Crystallography, X-Ray; D015497:HIV-1 / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D057927:Hydrophobic and Hydrophilic Interactions; D007128:Immunoglobulin Fragments; D008027:Light; D008744:Methylamines; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry*; D017510:Protein Folding; D012542:Scattering, Radiation; D053838:Scattering, Small Angle; D013056:Spectrophotometry, Ultraviolet; D014270:Trifluoroethanol; D014867:Water; D054322:tat Gene Products, Human Immunodeficiency Virus / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism
D016915:AIDS Vaccines; D007128:Immunoglobulin Fragments; D008744:Methylamines; D050505:Mutant Proteins; C421254:human immunodeficiency virus type 1 Tat vaccine; D054322:tat Gene Products, Human Immunodeficiency Virus; D014867:Water; D014270:Trifluoroethanol; C005855:trimethyloxamine