Isolation and Characterization of a Novel Haloacid Permease from Burkholderia cepacia MBA4

Abstract

ABSTRACT Burkholderia cepacia MBA4 is a bacterium that can utilize 2-haloacids as carbon and energy sources for growth. It has been proposed that dehalogenase-associated permease mediates the uptake of haloacid. In this paper, we report the first cloning and characterization of such a haloacid permease. The structural gene, designated deh4p , was found 353 bases downstream of the dehalogenase gene deh4a . Quantitative analysis of the expression of deh4p showed that it was induced by monochloroacetate (MCA), to a level similar to the MCA-induced level of deh4a . The nucleotide sequence of deh4p was determined, and an open reading frame of 1,656 bp encoding a putative peptide of 552 amino acids was identified. Deh4p has a putative molecular weight of 59,414 and an isoelectric point of 9.88. Deh4p has the signatures of sugar transport proteins and integral membrane proteins of the major facilitator superfamily. Uptake of [ 14 C]MCA into the cell was Deh4p dependent. Deh4p has apparent K m s of 5.5 and 8.9 μM and V max s of 9.1 and 23.1 nmol mg −1 min −1 for acetate and MCA, respectively. A mutant with a transposon-inactivated haloacid operon failed to grow on MCA even when deh4a was provided in trans .