Structurally homologous all beta-barrel proteins adopt different mechanisms of folding.
Acidic fibroblast growth factors from human (hFGF-1) and newt (nFGF-1) (Notopthalamus viridescens) are 16-kDa, all beta-sheet proteins with nearly identical three-dimensional structures. Guanidine hydrochloride (GdnHCl)-induced unfolding of hFGF-1 and nFGF-1 monitored by fluorescence and far-UV circular dichroism (CD) shows that the FGF-1 isoforms differ significantly in their thermodynamic stabilities. GdnHCl-induced unfolding of nFGF-1 follows a two-state (Native state to Denatured state(s)) mechanism without detectable intermediate(s). By contrast, unfolding of hFGF-1 monitored by fluorescence, far-UV circular dichroism, size-exclusion chromatography, and NMR spectroscopy shows that the unfolding process is noncooperative and proceeds with the accumulation of stable intermediate(s) at 0.96 M GdnHCl. The intermediate (in hFGF-1) populated maximally at 0.96 M GdnHCl has molten globule-like properties and shows strong binding affinity to the hydrophobic dye, 1-Anilino-8-naphthalene sulfonate (ANS). Refolding kinetics of hFGF-1 and nFGF-1 monitored by stopped-flow fluorescence reveal that hFGF-1 and nFGF-1 adopts different folding mechanisms. The observed differences in the folding/unfolding mechanisms of nFGF-1 and hFGF-1 are proposed to be either due to differential stabilizing effects of the charged denaturant (Gdn(+) Cl(-)) on the intermediate state(s) and/or due to differences in the structural interactions stabilizing the native conformation(s) of the FGF-1 isoforms.
Srimathi
Thiagarajan
T
Department of Chemistry, National Tsing Hua University, Hsinchu, Taiwan.
Kumar
Thallampuranam Krishnaswamy S
TK
Kathir
Karuppanan Muthusamy
KM
Chi
Ya-Hui
YH
Srisailam
Sampath
S
Lin
Wann-Yin
WY
Chiu
Ing-Ming
IM
Yu
Chin
C
D000595:Amino Acid Sequence; D000818:Animals; D002850:Chromatography, Gel; D041961:Deuterium Exchange Measurement; D016220:Fibroblast Growth Factor 1 / Q000737:chemistry* / Q000145:classification*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009673:Notophthalmus viridescens; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D020033:Protein Isoforms / Q000737:chemistry* / Q000145:classification*; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D013050:Spectrometry, Fluorescence; D013329:Structure-Activity Relationship
D020033:Protein Isoforms; D016220:Fibroblast Growth Factor 1