Purification of alpha-amylase isoenzymes from Scytalidium thermophilum on a fluidized bed of alginate beads followed by concanavalin A-agarose column chromatography.
An alpha-amylase has been purified from the thermophilic fungus Scytalidium thermophilum. A ninefold purification was achieved in a single step using fluidized bed chromatography wherein alginate was used as the affinity matrix. There are at least two isoenzymes as shown by concanavalin A (Con A)-agarose column chromatography. The isoenzyme binding to Con A is stable for at least 3 h at 80 degrees C in the presence of calcium ions. The isoenzymes have similar molecular weights of around 45,000 Da as shown by SDS-PAGE analysis. The isoenzymes differ only slightly in their pH optima and temperature optima but the isoenzyme binding to Con A-agarose has slightly higher thermal stability.
Roy
I
I
Chemistry Department, Indian Institute of Technology, Hauz Khas, New Delhi, 110 016, India.
Sastry
M S
MS
Johri
B N
BN
Gupta
M N
MN
D000327:Adsorption; D000464:Alginates / Q000378:metabolism*; D001203:Ascomycota / Q000201:enzymology*; D002118:Calcium / Q000494:pharmacology; D002846:Chromatography, Affinity; D003208:Concanavalin A / Q000378:metabolism*; D004795:Enzyme Stability / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000302:isolation & purification / Q000378:metabolism; D008863:Microspheres; D011485:Protein Binding; D013696:Temperature; D013816:Thermodynamics; D000516:alpha-Amylases / Q000302:isolation & purification* / Q000378:metabolism*
D000464:Alginates; D007527:Isoenzymes; D003208:Concanavalin A; D000516:alpha-Amylases; D002118:Calcium