Conformational changes of bovine α-lactalbumin induced by adsorption on a hydrophobic interface are studied by fluorescence and circular dichroism spectroscopy. Adsorption of bovine α-lactalbumin on hydrophobic polystyrene nanospheres induces a non-native state of the protein, which is characterized by preserved secondary structure, lost tertiary structure, and release of calcium. This partially denatured state therefore resembles a molten globule state, which is an intermediate in the folding of bovine α-lactalbumin. Stopped-flow fluorescence spectroscopy reveals two kinetic phases during adsorption with rate constants k1 ∼ 50 s−1 and k2 ∼ 8 s−1. The rate of partial unfolding is remarkably fast and even faster than unfolding induced by the addition of 5.4 mguanidinium hydrochloride to native α-lactalbumin. The large unfolding rates exclude the possibility that unfolding of bovine α-lactalbumin to the intermediate state occurs before adsorption takes place. Stopped-flow fluorescence anisotropy experiments show that adsorption of bovine α-lactalbumin on polystyrene nanospheres occurs within the dead time (15 ms) of the experiment. This shows that the kinetic processes as determined by stopped-flow fluorescence spectroscopy are not affected by diffusion or association processes but are solely caused by unfolding of bovine α-lactalbumin induced by adsorption on the polystyrene surface. A scheme is presented that incorporates the results obtained and describes the adsorption of bovine α-lactalbumin.
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Published, JBC Papers in Press, January 8, 2002, DOI 10.1074/jbc.M106005200
This research was financially supported by Senter, The Hague, The Netherlands (IOP-IE 98004). The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
