Utilization of trihalogenated propanes by Agrobacterium radiobacter AD1 through heterologous expression of the haloalkane dehalogenase from Rhodococcus sp strain m15-3

Trihalogenated propanes are toxic and recalcitrant organic compounds. Attem pts to obtain pure bacterial cultures able to use these compounds as sole c arbon and energy sources were unsuccessful. Both the haloalkane dehalogenas e from Xanthobacter autotrophicus GJ10 (DhlA) and that from Rhodococcus sp, strain m15-3 (DhaA) were found to dehalogenate trihalopropanes to 2,3-diha logenated propanols, but the kinetic properties of the latter enzyme are mu ch better, Broad-host-range dehalogenase expression plasmids, based on RSF1 010 derivatives, were constructed with the haloalkane dehalogenase from Rho dococcus sp, strain m15-3 under the control of the heterologous promoters P -lac, P-dhlA, and P-trc. The resulting plasmids yielded functional expressi on in several gram-negative bacteria. A catabolic pathway for trihalopropan es was designed by introducing these broad-host-range dehalogenase expressi on plasmids into Agrobacterium radiobacter AD1, which has the ability to ut ilize dihalogenated propanols for growth. The recombinant strain AD1 (pTB3) , expressing the haloalkane dehalogenase gene under the control of the dhlA promoter, was able to utilize both 1,2,3-tribromopropane and 1,2-dibromo-3 -chloropropane as sole carbon sources. Moreover, increased expression of th e haloalkane dehalogenase resulted in elevated resistance to trihalopropane s.