Characterization of an α-amylase from sorghum (Sorghum bicolor) obtained under optimized conditions

Abstract

Sorghummalt α-amylase can compete with bacterial α-amylase in industrial applications, if sufficiently stable and produced in a large enough quantity. Conditions for maximal α-amylase production in sorghum malt and the physico-chemical properties of the α-amylase so produced are reported in this study. Sorghum grains were steeped in buffers with varying pH (4.0-8.0) for 24 h, at room temperature, and germinated for another 48h to obtain the green malt. The buffer that induced the highest quantity of α-amylase was chosen as the optimal pH and served as the medium for further steeping experiments conducted at different temperatures (10, 20, 30, 40, 50 and 60°C). The α-amylase activity in the extract was determined in order to obtain the optimumtemperature for α-amylase induction at this particular pH. For the purpose of comparison, the α-amylase produced at the optimumpH and temperaturewas purified to apparent homogeneity by a combination of ion-exchange and size-exclusion chromatography, and further characterized. Eight-fold higher α-amylase activity was induced in pH 6.5 buffer at 20°C compared withwater, the traditional steeping medium. The Km and Vmaxwere estimated to be 1.092±0.05mgmL-1 and 3516±1.981 units min-1, respectively. The activation energy of the purified amylase for starch hydrolysis was 6.2 kcalK-1mol-1. Chlorides of calcium and manganese served as good activators, whereas CuSO4 inhibited the enzyme with a 42% loss in activity at 312mM salt concentration. © 2012 The Institute of Brewing & Distilling.