In vivo, α-crystallin and other small heat-shock proteins (sHsps) act as molecular chaperones to prevent the precipitation of ‘substrate’ proteins under stress conditions through the formation of a soluble sHsp–substrate complex. Using a range of different salt conditions, the rate and extent of precipitation of reduced α-lactalbumin have been altered. The interaction of α-crystallin with reduced α-lactalbumin under these various salt conditions was then studied using a range of spectroscopic techniques. Under conditions of low salt, α-lactalbumin aggregates but does not precipitate. α-Crystallin is able to prevent this aggregation, initially by stabilization of a monomeric molten-globule species of α-lactalbumin. It is proposed that this stabilization occurs through weak transient interactions between α-crystallin and α-lactalbumin. Eventually a stable, soluble high-molecular-mass complex is formed between the two proteins. Thus it appears that a tendency for α-lactalbumin to aggregate (but not necessarily precipitate) is the essential requirement for α-crystallin–α-lactalbumin interaction. In other words, α-crystallin interacts with a non-aggregated form of the substrate to prevent aggregation. The rate of precipitation of α-lactalbumin is increased significantly in the presence of Na2SO4 compared with NaCl. However, in the former case, α-crystallin is unable to prevent this aggregation and precipitation except in the presence of a large excess of α-crystallin, i.e. at mass ratios more than 10 times greater than in the presence of NaCl. It is concluded that a kinetic competition exists between aggregation and interaction of unfolding proteins with α-crystallin.

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