Engineering a heavy atom derivative for the X-ray structure analysis of cyclodextrin glycosyltransferase

Based on a preliminary structural model of cyclodextrin glycosyltransferasefrom Bacillus circulans (EC 2.4.1.19 EC] ), Ser428 and Ser475 ofthe enzyme were mutated to cysteines in order to produce suitableheavy atom derivatives. Mutant Ser475 - Cys could not be expressedas protein. Mutant Ser428 - Cys was expressed in Escherichiacoli and purified. It crystallized isomorphously and gave riseto a mercury derivative that improved the electron density map.The structural results show that the new mercury-binding siteis in a pocket at the protein surface.