Amyloid-like fibril formation in an all beta-barrel protein involves the formation of partially structured intermediate(s).
In the present study, we demonstrate the thermal induced amyloid formation in a beta-barrel protein, such as the acidic fibroblast growth factor from Notopthalmus viridescens (nFGF-1). Fibril formation in nFGF-1 is observed to occur maximally at 65 degrees C. Electron microscope analysis of the thermal induced fibrils of nFGF-1 shows that they are filamentous with an average diameter of about 20 nm. X-ray diffraction analysis reveals that the thermal induced fibrils of nFGF-1 have a typical "cross-beta" structure with the beta-strands perpendicular to the fibril axis. By using a variety of biophysical techniques including multidimensional NMR, we demonstrate that fibril formation involves the formation of a partially structured intermediate(s) in the thermal unfolding pathway of the protein (nFGF-1). Results of the anilino-8-napthalene sulfonate binding experiments indicate that fibril formation occurs due to the coalescence of the protein (in the intermediate state(s)) through the solvent-exposed non-polar surface(s). In this study, we also demonstrate that organic osmolytes, such as proline, can efficiently prevent the thermal induced amyloid formation in nFGF-1. Proline is found to stabilize the native conformation of the protein. The addition, proline is observed to increase the cooperativity of the unfolding (native <--> denatured) reaction and consequently decrease the population of the "sticky" thermal equilibrium intermediate(s) responsible for the fibril formation.
Srisailam
Sampath
S
Department of Chemistry, National Tsing Hua University, National Tsing Hua University, Hsinchu 30043, Taiwan.
Wang
Han-Min
HM
Kumar
Thallampuranam Krishnaswamy S
TK
Rajalingam
Dakshinamurthy
D
Sivaraja
Vaithiyalingam
V
Sheu
Hwo-Shuenn
HS
Chang
Yen-Chung
YC
Yu
Chin
C
D000682:Amyloid / Q000096:biosynthesis* / Q000737:chemistry / Q000648:ultrastructure; D000818:Animals; D002942:Circular Dichroism; D016220:Fibroblast Growth Factor 1 / Q000737:chemistry / Q000378:metabolism*; D006358:Hot Temperature; D008854:Microscopy, Electron; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular; D011487:Protein Conformation; D011489:Protein Denaturation; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D012455:Salamandridae; D017550:Spectroscopy, Fourier Transform Infrared; D014961:X-Ray Diffraction
D000682:Amyloid; D011994:Recombinant Proteins; D016220:Fibroblast Growth Factor 1