Recombinant expression of a unique chloromuconolactone dehalogenase ClcF from Rhodococcus opacus 1CP and identification of catalytically relevant residues by mutational analysis

► Recombinant ClcF and variants thereof were made available for structural studies. ► A homology model for ClcF was provided on the basis of muconolactone isomerase. ► A mechanistic role of E27 in proton abstraction is proposed. ► The effect of N52 on product specificity suggests its function in substrate fixation.