The λ Gam Protein Inhibits RecBCD Binding to dsDNA Ends

Murphy, Kenan C.

doi:10.1016/j.jmb.2007.05.085

Cited by 45 publications

(43 citation statements)

References 29 publications

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“…Gam protects the phage linear dsDNA from degradation by inhibiting the host's RecBCD endonuclease (25,48). Although we have not assessed the presence of other RecBCD inhibitors (46), the absence of Gam from plasmids and ICEs could be due to the fact that no step of these elements' normal "life cycle" involves a linear dsDNA molecule.…”

Section: Discussionmentioning

confidence: 95%

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DNA-Damaging Agents Induce the RecA-Independent Homologous Recombination Functions of Integrating Conjugative Elements of the SXT/R391 Family

2013

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Integrating conjugative elements (ICEs) of the SXT/R391 family are major contributors to the spread of antibiotic resistance genes. These elements also catalyze their own diversity by promoting inter-ICE recombination through the action of the RecA-independent homologous recombination system that they encode. Here, we report that expression of this recombination system, which consists of the single-stranded DNA annealing protein Bet and the exonuclease Exo, is induced by DNA-damaging agents via ICE-encoded transcriptional regulators. We show that the bet and exo genes are part of a large polycistronic transcript that contains many conserved ICE genes that are not involved in the main integration/excision and conjugative transfer processes. We show that although the recombination genes are highly transcribed, their translation is subject to additional strong regulatory mechanisms. We also show that an ICE-encoded putative single-stranded DNA binding protein (Ssb) limits hybrid ICE formation. Finally, a thorough in silico analysis reveals that orthologues of Bet and Exo are widely distributed in bacterial strains belonging to very distantly related bacterial species and are carried by various mobile genetic elements. Phylogenetic analyses suggest that the annealing proteins and exonucleases that compose these systems sometimes have different evolutionary origins, underscoring the strong selective pressure to maintain the functionality of these unrelated cooperating proteins.

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Section: Discussionmentioning

confidence: 95%

“…1). The product of gam protects the phage's linear double-stranded DNA (dsDNA) by inhibiting the ability of RecBCD to bind ds-DNA ends (25). No homologue of gam exists in SXT/R391 ICEs or IncA/C plasmids; instead, a gene encoding a putative singlestranded DNA-binding protein (ssb) is found upstream of bet.…”

mentioning

confidence: 99%

DNA-Damaging Agents Induce the RecA-Independent Homologous Recombination Functions of Integrating Conjugative Elements of the SXT/R391 Family

2013

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“…The Red genes are expressed together with the λ gam gene under a temperature-inducible promoter for the efficient induction and DNA manipulation in bacteria. Gam is a natural inhibitor of the E. coli RecBCD exonuclease, which rapidly degrades dsDNA invading into bacteria (Karu et al, 1975;Murphy, 1991Murphy, , 1998Murphy, , 2007Yu et al, 2000). While the Red recombination system is expressed, a linear DNA fragment with 40-50 bp homologous flanking regions is inserted into to the selected target sequence by Exo and Beta, whereas Gam blocks the RecBCD enzyme from degrading dsDNA ends.…”

Section: Site-directed Mutagenesis Of Viral Bacterial Artificial Chromentioning

confidence: 99%

“…Moreover, the necessity for virus replication was determined for 162 individual hCMV genes (Dunn et al, 2003). mCMV as an important animal model for hCMV pathogenesis was the first herpesvirus genome to be cloned as an infectious BAC (Messerle et al, 1997) which has been used for in vitro and in vivo mCMV studies (e.g., Wagner et al, 1999;Cicin-Sain et al, 2003, 2007Menard et al, 2003;Schnee et al, 2006). BAC-clones have also been constructed for the genomes of rhesus CMV (rhCMV; Chang & Barry, 2003;Lilja et al, 2008;Rue et al, 2004) and guinea-pig CMV (gpCMV; Crumpler et al, 2009;McGregor & Schleiss, 2001a;Schleiss, 2008).…”

Section: Functional Mutagenesis Of Specific Viral Bacsmentioning

confidence: 99%

Bacterial Genetics of Large Mammalian DNA Viruses: Bacterial Artificial Chromosomes as a Prerequisite for Efficiently Studying Viral DNA Replication and Functions

Wussow¹,

Spieckermann²,

Brunnemann³

et al. 2011

DNA Replication-Current Advances

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“…[65] The Gam protein inhibits exonuclease RecBCD binding to dsDNA ends. [64,66] The Rac prophage was recently used for large DNA assembly in E. coli. [67] The RecET recombination system of the Rac prophage is functionally analogous to the Red system of the bacteriophage lambda (RecE and RecT are analogous to Exo and Beta, respectively).…”

Section: Introductionmentioning

confidence: 99%

High molecular weight DNA assembly in vivo for synthetic biology applications

Juhas

Ajioka

2016

Critical Reviews in Biotechnology

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DNA assembly is the key technology of the emerging interdisciplinary field of synthetic biology. While the assembly of smaller DNA fragments is usually performed in vitro, high molecular weight DNA molecules are assembled in vivo via homologous recombination in the host cell. Escherichia coli, Bacillus subtilis and Saccharomyces cerevisiae are the main hosts used for DNA assembly in vivo. Progress in DNA assembly over the last few years has paved the way for the construction of whole genomes. This review provides an update on recent synthetic biology advances with particular emphasis on high molecular weight DNA assembly in vivo in E. coli, B. subtilis and S. cerevisiae. Special attention is paid to the assembly of whole genomes, such as those of the first synthetic cell, synthetic yeast and minimal genomes.

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The λ Gam Protein Inhibits RecBCD Binding to dsDNA Ends

Cited by 45 publications

References 29 publications

DNA-Damaging Agents Induce the RecA-Independent Homologous Recombination Functions of Integrating Conjugative Elements of the SXT/R391 Family

DNA-Damaging Agents Induce the RecA-Independent Homologous Recombination Functions of Integrating Conjugative Elements of the SXT/R391 Family

Bacterial Genetics of Large Mammalian DNA Viruses: Bacterial Artificial Chromosomes as a Prerequisite for Efficiently Studying Viral DNA Replication and Functions

High molecular weight DNA assembly in vivo for synthetic biology applications

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