On the edge of the denaturation process: application of X-ray diffraction to barnase and lysozyme cross-linked crystals with denaturants in molar concentrations.
Structural data about the early step of protein denaturation were obtained from cross-linked crystals for two small proteins: barnase and lysozyme. Several denaturant agents like urea, bromoethanol or thiourea were used at increasing concentrations up to a limit leading to crystal disruption (>or=2 to 6 M). Before the complete destruction of the crystal order started, specific binding sites were observed at the protein surfaces, an indication that the preliminary step of denaturation is the disproportion of intermolecular polar bonds to the benefit of the agent "parasiting" the surface. The analysis of the thermal factors first agree with a stabilization effect at low or moderate concentration of denaturants rapidly followed by a destabilization at specific weak points when the number of sites increase (overflooding effect).
Salem
Michèle
M
Université René Descartes, Faculté de pharmacie, Laboratoire de cristallographie et RMN biologiques (UMR-8015, CNRS), 4 av. de l'Observatoire 75270 Paris Cedex 06, France.
Mauguen
Yves
Y
Prangé
Thierry
T
D000818:Animals; D001407:Bacillus / Q000201:enzymology; D001426:Bacterial Proteins; D002645:Chickens; D003432:Cross-Linking Reagents / Q000737:chemistry; D003460:Crystallization; D018360:Crystallography, X-Ray; D000431:Ethanol / Q000031:analogs & derivatives; D009113:Muramidase / Q000737:chemistry* / Q000378:metabolism; D010766:Phosphorylation; D011487:Protein Conformation; D011489:Protein Denaturation; D012260:Ribonucleases / Q000737:chemistry* / Q000378:metabolism*; D013696:Temperature; D013890:Thiourea; D014508:Urea
D001426:Bacterial Proteins; D003432:Cross-Linking Reagents; D000431:Ethanol; D014508:Urea; D012260:Ribonucleases; C019760:Bacillus amyloliquefaciens ribonuclease; D009113:Muramidase; D013890:Thiourea