%0 Journal Article
%T DNA phosphorothioation in Streptomyces lividans: mutational analysis of the dnd locus
%A Tiegang Xu
%A Jingdan Liang
%A Shi Chen
%A Lianrong Wang
%A Xinyi He
%A Delin You
%A Zhijun Wang
%A Aiying Li
%A Zhongli Xu
%A Xiufen Zhou
%A Zixin Deng
%J BMC Microbiology
%D 2009
%I BioMed Central
%R 10.1186/1471-2180-9-41
%X A 6,665-bp DNA region carrying just five ORFs (dndA-E) was defined as the sole determinant for modification of the DNA backbone in S. lividans to form phosphorothioate. This provides a diagnostically reliable and easily assayable Dnd (DNA degradation) phenotype. While dndA is clearly transcribed independently, dndB-E constitute an operon, as revealed by RT-PCR analysis. An efficient mutation-integration-complementation system was developed to allow for detailed functional analysis of these dnd genes. The Dnd- phenotype caused by specific in-frame deletion of the dndA, C, D, and E genes or the enhanced Dnd phenotype resulting from in-frame deletion of dndB could be restored by expression vectors carrying the corresponding dnd genes. Interestingly, overdosage of DndC or DndD, but not other Dnd proteins, in vivo was found to be detrimental to cell viability.DNA phosphorothioation is a multi-enzymatic and highly coordinated process controlled by five dnd genes. Overexpression of some proteins in vivo prevented growth of host strain, suggesting that expression of the gene cluster is strictly regulated in the native host.Most of the commonly found structural changes in DNA are due to methylation of selected bases. In some viral DNAs, certain bases may be hydroxymethylated or glucosylated [1-3]. Altered or unusual bases in DNA molecules often have significant physiological implications, such as DNA replication control, gene regulation, or protection of the respective organisms from invasion by foreign DNA [4].In contrast to other types of DNA modification, S, lividans has a site-specific and stereo-selective sulfur modification on the DNA backbone termed phosphorothioation [5-7]. This sulfur modification occurs specifically between two guanine nucleotides in S.lividans [6,8]. The sulfur-modified DNA suffers double-stranded cleavage at the modification sites during normal and pulsed-field gel electrophoresis [6,9-13]. The Dnd phenotype was proven to be a peracid-mediated, o
%U http://www.biomedcentral.com/1471-2180/9/41