1 research outputs found
Heterologous Production and Purification of a Functional Chloroform Reductive Dehalogenase
Reductive
dehalogenases (RDases) are key enzymes involved in the
respiratory process of anaerobic organohalide respiring bacteria (ORB).
Heterologous expression of respiratory RDases is desirable for structural
and functional studies; however, there are few reports of successful
expression of these enzymes. <i>Dehalobacter</i> sp. strain
UNSWDHB is an ORB, whose preferred electron acceptor is chloroform.
This study describes efforts to express recombinant reductive dehalogenase
(TmrA), derived from UNSW DHB, using the heterologous hosts <i>Escherichia coli</i> and <i>Bacillus megaterium</i>. Here, we report the recombinant expression of soluble and functional
TmrA, using <i>B. megaterium</i> as an expression host under
a xylose-inducible promoter. Successful incorporation of iron–sulfur
clusters and a corrinoid cofactor was demonstrated using UV–vis
spectroscopic analyses. <i>In vitro</i> dehalogenation of
chloroform using purified recombinant TmrA was demonstrated. This
is the first known report of heterologous expression and purification
of a respiratory reductive dehalogenase from an obligate organohalide
respiring bacterium