STRUCTURE OF CYCLODEXTRIN GLYCOSYLTRANSFERASE COMPLEXED WITH A MALTONONAOSE INHIBITOR AT 2.6 ANGSTROM RESOLUTION - IMPLICATIONS FOR PRODUCTSPECIFICITY

Crystals of the Y195F mutant of cyclodextrin glycosyltransferase from Bacillus circulans strain 251 were subjected to a double soaking proce dure, in which they were first soaked in a solution containing the inh ibitor acarbose and subsequently in a solution containing maltohexaose . The refined structure of the resulting protein-carbohydrate complex has final crystallographic and free R-factors for data in the 8-2.6 An gstrom resolution range of 15.0% and 21.5%, respectively, and reveals that a new inhibitor, composed of nine saccharide residues, is bound i n the active site. The first four residues correspond to acarbose and occupy the same subsites near the catalytic residues as observed in th e previously reported acarbose-enzyme complex [Strokopytov et al. (199 5) Biochemistry 34, 2234-2240]. An oligosaccharide consisting of five glucose residues has been coupled to the nonreducing end of acarbose. At the fifth residue the polysaccharide chain makes a sharp turn, allo wing it to interact with residues Tyr89, Phe195, and Asn193 and a flex ible loop formed by residues 145-148. On the basis of the refined mode l of the complex an explanation is given for the product specificity o f CGTases.