In vitro analysis of aggregation-disaggregation of the folding intermediate of Bacillus subtilis α-amylase.

Anne Colomer-Pallas (1) , Marie-Françoise Petit-Glatron (1) , Regis Chambert (1)

1 IJM (UMR_7592) - Institut Jacques Monod

The refolding intermediate of Bacillus subtilis α-amylase is prone to aggregate at 37°C and pH 7 when the protein concentration is relatively high (≥ 1µM). Low concentrations of 2,2,2 trifluoroethanol greatly increased the rate of aggregation. Aggregation made the folding intermediate resistant to proteases and there was kinetic competition between aggregation and proteolytic degradation. Analysis by Fourier transform infrared spectroscopy indicated that the secondary structure of the refolding intermediate is sightly different under soluble or aggregated states. Aggregates were readily solubilized by guanidium chloride (D1/2 = 1.25 M), but disaggregation was slow when aggregates were resuspended in solutions of various foreign native proteins under physiological conditions of pH and temperature. This destabilizing effect resulting from protein-protein interactions may make the aggregation process reversible in vivo.

Mots clés: alpha-amylase; Bacillus subtilis; protein secretion; aggregation-disaggregation; folding intermediate

Domaines: Biologie cellulaire

Anne Colomer-Pallas, Marie-Françoise Petit-Glatron, Regis Chambert. In vitro analysis of aggregation-disaggregation of the folding intermediate of Bacillus subtilis α-amylase.. Journal of Biological Physics and Chemistry, 2002, 2, pp.101-107. ⟨hal-00019349⟩