Ligand-induced conformational changes in villin, a calcium-controlled actin-modulating protein.
The physical structure of villin, a Ca2+-modulated regulator protein of actin filament organization, has been studied in the absence and presence of Ca2+ using analytical ultracentrifugation, gel chromatography, ultraviolet difference spectroscopy, and circular dichroism. Villin exhibits an intrinsic sedimentation coefficient, S020, w, of 5.0 s and an apparent Stokes radius, Rs, of 44 A in EGTA-containing buffer. In the presence of greater than 20 microM Ca2+, villin shows a S020, w of 4.1 s and Rs of 49 A, indicative of a conformational change in the protein. No significant changes in the partial specific volume (0.73) of villin are observed in the presence of Ca2+, and sedimentation equilibrium studies demonstrates that the effects of Ca2+ are not due to a change in the molecular mass (95,000 daltons). The combined hydrodynamic data suggest that villin is an asymmetric molecule with an axial ratio of 4.5:1, based on a prolate ellipsoid model at 0.5 g/g of hydration, corresponding to a maximum length of 84 A. The presence of Ca2+ changes the shape to a more asymmetric molecule with an axial ratio of 8:1 and a maximum length of 123 A. Since the large proteolytic fragment, villin core, does not exhibit the strong structural change of villin, an important function of the small villin headpiece domain in the observed conformational change of intact villin is suggested. The results indicate new aspects of the function of villin as cross-linker in microvillus core filament bundles and the disintegration of these structures in the presence of calcium.
D000199:Actins / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002352:Carrier Proteins / Q000302:isolation & purification / Q000378:metabolism*; D002645:Chickens; D002942:Circular Dichroism; D007421:Intestine, Small / Q000378:metabolism; D008024:Ligands; D008840:Microfilament Proteins; D011487:Protein Conformation; D013056:Spectrophotometry, Ultraviolet
D000199:Actins; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D008024:Ligands; D008840:Microfilament Proteins; C026615:villin; D002118:Calcium