Abstract

Oyefuga O. H, Adeyanju M. M, Adebawo. O. O. and Agboola F. K

-amylase (EC 3.2.1.2, -1,4–D-glucan maltohydrolase) was isolated and purified from the nodes of sugar cane by using ammonium sulphate precipitation, acid-treatment, gel filtration on Sephadex G-75 and ion exchange chromatography on DEAE-Cellulose. Purity was ascertained by the presence of a single band of protein on polyacrylamide gel electrophoresis under non-denaturing conditions. The specific activity was 4.68 unit.mg-1 of protein and recovery of 15.67%. The Km value of the enzyme for starch as substrate was 3.20% while its Vmax was 1.11units.min-1 .ml-1 . The apparent molecular weight was estimated by gel filtration on a Sephadex G-200 column to be 156,000 Da. The subunit molecular weight was found to be 154,000 Da by sodium dodecyl sulphate polyacrylamide gel electrophoresis. This suggests that the enzyme exists in a monomeric form. The optimum pH for the activity of the enzyme was pH 5.5, while its optimum temperature was 60°C. The hydrolysate of the action of the enzyme showed maltose as the main product of hydrolysis on thin layer chromatography.