Relationship between molecular states (conformation and orientation) and activities of α-amylase adsorbed on ultrafine silica particles

Abstract

Bacillus subtilis α-amylase, which contains a relatively large amount of α-helix, was adsorbed on two types of ultrafine silica particles (silica-1 and-2, average diameter 15 nm) under various conditions. The changes in circular dichroism (CD) spectra of α-amylase upon adsorption were measured, and the extent of conformational changes was estimated from the reduction in α-helix content. In additions the activities of adsorbed α-amylase were measured at pH 5.2 using corn starch andp-nitrophenylbenzyl α-maltopentaoside (BG5P). In the ultrafine silica-2 particles, the extent of both activity reductions and conformational changes upon adsorption was much larger than that in the ultrafine silica-1 particles and increased with decreasing pH and amount of adsorption. The extent of activity reductions correlated closely with the conformational changes. On the other hand, the effect of reduction in α-amylase activity upon adsorption measured by BG5P was smaller than that measured by starch, indicating that the lack of accessibility of the active site to a large substrate also reduces the activity of adsorbed α-amylase. However, the effects of particle type and adsorption conditions on the extent of activity reductions by the accessibility resistance were small. Therefore, variation of the activity of adsorbed α-amylase is mainly attributable to the extent of conformational changes upon adsorption. Based on these results, a procedure to prepare adsorbed α-amylase with high activity was investigated.