In order to investigate the role of different residues of Methylophilus sp. strain DM11 dichloromethane dehalogenase for substrate binding, glutathione affinity, and catalytic activity, site-directed mutagenesis studies of the gene encoding the enzyme were carried out. The conserved tryptophane residue at 103 region was respectively substituted by phenylalanine, valine or asparagine. The conserved arginine residue at 109 region was substituted by leucine. The conserved tryptophane residue at 117 region was respectively substituted by tyrosine or phenylalanine. Six mutant enzymes were produced. Among them three possess lower activities, other three do not possess activity. The properties of the mutant enzyme W117Y are very different from wild-type enzyme.