Purification and characterization of a thiol amylase over produced by a non-cereal non-leguminous plant, Tinospora cordifolia.

A 43kDa α-amylase was purified from Tinospora cordifolia by glycogen precipitation, ammonium sulfate precipitation, gel filtration chromatography, and HPGPLC. The enzyme was optimally active in pH 6.0 at 60°C and had specific activity of 546.2U/mg of protein. Activity was stable in the pH range of 4-7 and at temperatures up to 60°C. PCMB, iodoacetic acid, iodoacetamide, DTNB, and heavy metal ions Hg(2+)>Ag(+)>Cd(2+) inhibited enzyme activity while Ca(2+) improved both activity and thermostability. The enzyme was a thiol amylase (3 SH group/mole) and DTNB inhibition of activity was released by cysteine. N-terminal sequence of the enzyme had poor similarity (12-24%) with those of plant and microbial amylases. The enzyme was equally active on soluble starch and amylopectin and released maltose as the major end product.