Sequence- and activity-based screening of microbial genomes for novel dehalogenases.
Bottom Line: Reliable sequence determinants are necessary to harness genome sequencing-efforts for accelerating the discovery of novel dehalogenases with improved or modified activities.In an attempt to extract dehalogenase sequence fingerprints, 103 uncharacterized potential dehalogenase candidates belonging to the α/β hydrolase (ABH) and haloacid dehalogenase-like hydrolase (HAD) superfamilies were screened for dehalogenase, esterase and phosphatase activity.Four new L-2-haloacid dehalogenases from the HAD superfamily were found to hydrolyse fluoroacetate, an activity never previously ascribed to enzymes in this superfamily.
Affiliation: Department of Biochemistry, University of Toronto, 1 King's College Circle, Toronto, Ontario M5S 1A8, Canada.Show MeSH
Mentions: For the identification of haloalkane dehalogenases in the ABH superfamily, the Asp catalytic nucleophile serves as an important selection criterion because it is fully conserved and essential for dehalogenation (Fig. 2). The pair of primary halide‐binding residues can act as an additional haloalkane dehalogenase indicator. It comprises: a Trp at the nucleophile + 1 position; and either a Trp that aligns with Trp175 of DhlA (a type I haloalkane dehalogenase) or an Asn equivalent to Asn28 of SAV4779 (a type II haloalkane dehalogenase) within the upstream His‐Gly‐Asn‐Pro motif.
Affiliation: Department of Biochemistry, University of Toronto, 1 King's College Circle, Toronto, Ontario M5S 1A8, Canada.