EXPRESSING DEINOCOCCUS RADIODURANS RECD IN ESCHERICHIA COLI: PHENOTYPIC EFFECTS IN RECBCD(-) AND RECD(-) CELLS

Abstract

The RecD helicase is a member of the RecBCD complex, which is essential for repair of double-stranded breaks (DSBs) in DNA via homologous recombination in Escherichia coli. The microbe Deinococcus radiodurans is capable of fixing high amounts of DSBs, owing to an efficient repair system. However, Deinococcus does not contain any recB or recC genes; only a RecD-like helicase has been observed.

In Escherichia coli strains mutant for the native RecD subunit, the D. radiodurans RecD cannot restore nuclease activity when infected with T4 gene2- bacteriophage. Cell viability tests and mitomycin C exposure of RecBC(+)D(-) strains show no adverse effects from the Dr RecD. A negative phenotype was encountered with strains lacking RecBCD and expressing D. radiodurans RecD protein. Microscopy studies of RecBCD(-) E. coli expressing the D. radiodurans RecD helicase show long cellular structures termed filaments. The Dr RecD protein may be binding to and disrupting the replication fork.