Single-step purification of a recombinant thermostable alpha-amylase aftersolubilization of the enzyme from insoluble aggregates

The expression of the gene encoding a thermostable a-amylase (EC 3.2.1.1) ( optimal activity at 100 degrees C) from the hyperthermophilic archaeon Pyro coccus woesei in the mesophilic hosts Escherichia coli and Halomonas elonga ta resulted in the formation of insoluble aggregates. More than 85% of the recombinant enzyme was present within the cells as insoluble but catalytica lly active aggregates. The recombinant alpha-amylase was purified to homoge neity in a single step by hydrophobic interaction chromatography on a pheny l superose column after solubilization of the enzyme under nondenaturing co nditions. The enzyme was purified 258-fold with a final yield of 54%. (C) 2 000 Published by Elsevier Science B.V. All rights reserved.