CRYSTALLOGRAPHIC ANALYSIS OF THE CATALYTIC MECHANISM OF HALOALKANE DEHALOGENASE

Crystal structures of haloalkane dehalogenase were determined in the p resence of the substrate 1,2-dichloroethane. At pH 5 and 4-degrees-C, substrate is bound in the active site without being converted; warming to room temperature causes the substrate's carbon-chlorine bond to be broken, producing a chloride ion with concomitant alkylation of the a ctive-site residue Asp124. At pH 6 and room temperature the alkylated enzyme is hydrolysed by a water molecule activated by the His289-Asp26 0 pair in the active site. These results show that catalysis by the de halogenase proceeds by a two-step mechanism involving an ester interme diate covalently bound at Asp124.