Guanidinium chloride- and urea-induced unfolding of FprA, a mycobacterium NADPH-ferredoxin reductase: stabilization of an apo-protein by GdmCl.
The guanidinium chloride- and urea-induced unfolding of FprA, a mycobacterium NADPH-ferredoxin reductase, was examined in detail using multiple spectroscopic techniques, enzyme activity measurements and size exclusion chromatography. The equilibrium unfolding of FprA by urea is a cooperative process where no stabilization of any partially folded intermediate of protein is observed. In comparison, the unfolding of FprA by guanidinium chloride proceeds through intermediates that are stabilized by interaction of protein with guanidinium chloride. In the presence of low concentrations of guanidinium chloride the protein undergoes compaction of the native conformation; this is due to optimization of charge in the native protein caused by electrostatic shielding by the guanidinium cation of charges on the polar groups located on the protein side chains. At a guanidinium chloride concentration of about 0.8 m, stabilization of apo-protein was observed. The stabilization of apo-FprA by guanidinium chloride is probably the result of direct binding of the Gdm+ cation to protein. The results presented here suggest that the difference between the urea- and guanidinium chloride-induced unfolding of FprA could be due to electrostatic interactions stabilizating the native conformation of this protein.
Shukla
Nidhi
N
Division of Molecular and Structural Biology, Central Drug Research Institute, Lucknow, India.
Bhatt
Anant Narayan
AN
Aliverti
Alessandro
A
Zanetti
Giuliana
G
Bhakuni
Vinod
V
D002942:Circular Dichroism; D004795:Enzyme Stability; D019791:Guanidine / Q000737:chemistry*; D009247:NADH, NADPH Oxidoreductases / Q000737:chemistry* / Q000378:metabolism; D011487:Protein Conformation; D011489:Protein Denaturation; D013050:Spectrometry, Fluorescence; D055672:Static Electricity; D014508:Urea / Q000737:chemistry*
D014508:Urea; C464438:FprA protein, M tuberculosis; D009247:NADH, NADPH Oxidoreductases; D019791:Guanidine