Engineering of Cyclodextrin Product Specificity and pH Optima of the Thermostable Cyclodextrin Glycosyltransferase from

The product specificity and pH optimum of the ther-mostable cyclodextrin glycosyltransferase (CGTase) from Thermoanaerobacterium thermosulfurigenes EM1 was engineered using a combination of x-ray crystallog-raphy and site-directed mutagenesis. Previously, a crys-tal soaking experiment with the Bacillus circulans strain 251 b-CGTase had revealed a maltononaose inhib-itor bound to the enzyme in an extended conformation. An identical experiment with the CGTase from T. ther-mosulfurigenes EM1 resulted in a 2.6-Å resolution x-ray structure of a complex with a maltohexaose inhibitor, bound in a different conformation. We hypothesize that the new maltohexaose conformation is related to the enhanced a-cyclodextrin production of the CGTase. The detailed structural information subsequently al