Crystallization of type I chloramphenicol acetyltransferase: An approach based on the concept of ionic strength reducers

Abstract

Chloramphenicol acetyltransferase (CAT) is responsible for bacterial resistance to chloramphenicol. It catalyzes inactivation of the antibiotic by acetyl-group transfer from acetyl CoA to one or both hydroxyl groups of chloramphenicol. Type I CAT possesses some unique properties which are not observed in other CAT variants. Type I CAT overexpressed in Escherichia coli was purified and crystals with a resolution limit of 2.22 Å have been obtained using a novel procedure which is based on the concept of 'ionic strength reducers'. The crystals have the symmetry of space group P1 and unit-cell parameters a = 96.46, b = 113.86, c= 114.21 Å, α= 119.9, β= 94.1, γ = 98.6°. These dimensions are consistent with four to six trimers per unit cell, corresponding to a solvent fraction ranging from 65 to 47%.