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Purification and properties of Rhizobial DehL expressed in Escherichia coli
Abstract
The Rhizobium sp. DehL was produced by heterologous expression of the cloned gene in Escherichia coli. DehL enzyme was purified to homogeneity and characterized. The molecular weights were
estimated to be 61 and 31 kDa by gel filtration and DS-polyacrylamide gel electrophoresis (SDSPAGE), respectively, suggesting that the enzyme is a dimer. The purified enzyme was specific to the Lisomer
monochloropropionate (L-2CP) and dichloroacetate (DCA). This protein was not able to act on 2,2-dichloropropionate (2,2DCP) and trichloroacetate (TCA). The estimated kinetic data indicated that
this enzyme has high affinity to its specific substrates. By searching protein amino acid sequence database, the predicted amino acid sequence of DehL showed a high level of homology to those Lspecific
monochloropropionate (D,L-2CP) dehalogenase of Rhizobium sp. NHG3 with 53% sequence identity. The amino acid sequence of DehL showed low level sequence identity to those of Class 1D dehalogenases, suggesting DehL from Rhizobium sp. may belong to different group of dehalogenase classification preferably Class 1L dehalogenase.
estimated to be 61 and 31 kDa by gel filtration and DS-polyacrylamide gel electrophoresis (SDSPAGE), respectively, suggesting that the enzyme is a dimer. The purified enzyme was specific to the Lisomer
monochloropropionate (L-2CP) and dichloroacetate (DCA). This protein was not able to act on 2,2-dichloropropionate (2,2DCP) and trichloroacetate (TCA). The estimated kinetic data indicated that
this enzyme has high affinity to its specific substrates. By searching protein amino acid sequence database, the predicted amino acid sequence of DehL showed a high level of homology to those Lspecific
monochloropropionate (D,L-2CP) dehalogenase of Rhizobium sp. NHG3 with 53% sequence identity. The amino acid sequence of DehL showed low level sequence identity to those of Class 1D dehalogenases, suggesting DehL from Rhizobium sp. may belong to different group of dehalogenase classification preferably Class 1L dehalogenase.
