DOMAIN-E OF BACILLUS-MACERANS CYCLODEXTRIN GLUCANOTRANSFERASE - AN INDEPENDENT STARCH-BINDING DOMAIN

The starch-binding domains of glucoamylase I (SBD of GA-I) from Asperg illus awamori and of cyclodextrin glucanotransferase (domain E of CGTa se) from Bacillus macerans were fused to the C-terminus of beta-galact osidase (beta-gal). The majority of the fusion proteins produced in Es cherichia coli were found as inclusion bodies. Active fusion proteins were purified by partial solubilization of the inclusion bodies with 2 M urea followed by affinity chromatography. Adsorption isotherms of p urified fusion proteins on corn starch and cross-linked amylose were g enerated. The beta-gal fusion proteins had similar affinities for cros s-linked amylose and corn starch but significantly different saturatio n capacities on corn starch. The adsorption and elution data from the potato starch column as well as the adsorption isotherms of beta-gal-d omain E fusion protein (BDE109) on corn starch and cross-linked amylos e demonstrated that domain E of CGTase is an independent domain, which retained its starch-binding activity when separated from the other fo ur (A-D) domains in CGTase. (C) 1995 John Wiley & Sons, Inc.