Identification and disruption analysis of the recN gene in the extremely radioresistant bacterium Deinococcus radiodurans

We isolated a radiosensitive mutant strain, KR4128, from a wild-type strain of Deinococcus radiodurans, which is known as a extremely radioresistant b acterium. The gene that restore the defect of the mutant in DNA repair was cloned and it turned out to be the homolog of the recN gene of Escherichia coli, The recN gene encoded a protein of 58 kDa, and, in its N-terminal reg ion, a potential ATP binding domain was conserved as expected for a prokary otic RecN protein. An analysis of sequence of the mutant recN gene revealed a G:C to T:A transversion near the 3' end of the coding region. This alter ation causes an ochre mutation, and results in the truncation of 47 amino a cids from the C-terminal region of the RecN protein. The null mutant of rec N gene was constructed by insertional mutagenesis, and it showed substantia l sensitivities to various types of DNA damaging agents, indicating that a single defect in the recN gene can directly affect the DNA damage resistant phenotype in D. radiodurans. The recN locus of KR4128 was also disrupted a nd the disruptant indicated the sensitivity that was indistinguishable from its progenitor. The result indicate that the transversion in the recN gene of KR4128 cells causes a complete loss of function of the RecN protein and thus the C-terminal region of the RecN protein includes domain essential t o its function. (C) 1999 Elsevier Science B.V. All rights reserved.