Assessment of the stability and unfolding pathways of azurin from Pseudomonas aeruginosa through the combination of denaturating osmolytes.
The denaturating osmolytes urea and guanidine hydrochloride (GuHCl) interact differently with azurin from Pseudomonas aeruginosa. At room temperature, even high concentrations of urea were unable to unfold the metalloprotein: instead a "partially unfolded" intermediate state is formed. In contrast, the protein unfolded state was formed at GuHCl, concentrations above approximately 3M and the unfolded state subsequently undergoes an irreversible reaction, which was studied employing absorbance and fluorescence spectroscopic techniques. The partially unfolded protein formed in the presence of urea completely unfolds upon adding GuHCl and we report a kinetic and thermodynamic study to characterize the process, taking into consideration that the unfolded state undergoes an irreversible reaction.
University of Puerto Rico, Department of Chemistry, Río Piedras Campus, P.O. Box 23346, San Juan, PR 00931-3346, USA.
Qing
Q
Huang
Edwin
E
Quiñones
D001400:Azurin / Q000737:chemistry*; D019791:Guanidine / Q000737:chemistry*; D007700:Kinetics; D011489:Protein Denaturation; D017510:Protein Folding; D011550:Pseudomonas aeruginosa / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet; D014508:Urea / Q000737:chemistry*
D001400:Azurin; D014508:Urea; D019791:Guanidine