Initial hydrophobic collapse in the folding of barstar

Abstract

TWO models are commonly used to describe the poorly understood earliest steps of protein folding. The framework model^1-3 stresses very early formation of nascent secondary structures, which coalesce into a compact, molten, globule-like form^4,5 from which tertiary structure slowly develops^6,7. The hydrophobic collapse model^8-10 gives overriding precedence to a nonspecific collapse of the polypeptide chain which facilitates subsequent formation of specific secondary and tertiary structure^11,12. Here we report our analysis of the earliest observable events of the major folding pathway of barstar, a small protein. We compare the kinetics of folding using circular dichroism at 222 nm and 270 nm, intrinsic trypto-phan fluorescence, fluorescence of the hydrophobic dye 8-anilino-1-naphthalene-sulphonic acid on binding, and restoration of trypto-phan-dansyl fluorescence energy transfer as structure-monitoring probes. We show that the polypeptide chain rapidly collapses (within 4 ms) to a compact globule with a solvent-accessible hydro-phobic core⁶, but with no optically active secondary or tertiary structure. Thus the earliest event of the major folding pathway of barstar is a nonspecific hydrophobic collapse that does not involve concomitant secondary structure formation.