Escherichia coli DNA helicase I catalyzes a site- and strand-specific nicking reaction at the F plasmid oriT

Matson, Steven W.; Morton, Brian

doi:10.1016/s0021-9258(18)98540-6

Cited by 86 publications

(14 citation statements)

References 29 publications

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“…Discovery of the proteins responsible for this relaxation had to wait for almost two decades (9). Biochemical characterization of the covalent interaction between the relaxase and its cognate nic site was first reported for the TraI relaxase of IncP plasmid RP4 (10), and similar features were soon found for the relaxases of other conjugative and mobilizable plasmids (11)(12)(13)(14). Relaxases were then related through a set of three conserved motifs to other ssDNA endonucleases involved in DNA replication and transposition (15,16), which defined the HUH superfamily of site-specific ssDNA endonucleases.…”

Section: Accepted Manuscriptmentioning

confidence: 99%

The secret life of conjugative relaxases

Guzmán-Herrador

Llosa

2019

Plasmid

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Conjugative relaxases are well-characterized proteins responsible for the siteand strand-specific endonucleolytic cleavage and strand transfer reactions taking place at the start and end of the conjugative DNA transfer process. Most of the relaxases characterized biochemically and structurally belong to the HUH family of endonucleases. However, an increasing number of new families of relaxases are revealing a variety of protein folds and catalytic alternatives to accomplish conjugative DNA processing. Relaxases show high specificity for their cognate target DNA sequences, but several recent reports underscore the importance of their activity on secondary targets, leading to widespread mobilization of plasmids containing an oriTlike sequence. Some relaxases perform other functions associated with their nicking and strand transfer ability, such as catalyzing site-specific recombination or initiation of plasmid replication. They perform these roles in the absence of conjugation, and the validation of these functions in several systems strongly suggest that they are not mere artifactual laboratory observations. Other unexpected roles recently assigned to relaxases include controlling plasmid copy number and promoting retrotransposition.Their capacity to mediate promiscuous mobilization and genetic reorganizations can be exploited for a number of imaginative biotechnological applications. Overall, there is increasing evidence that conjugative relaxases are not only key enzymes for horizontal gene transfer, but may have been adapted to perform other roles which contribute to prokaryotic genetic plasticity. Relaxed target specificity may be key to this versatility.

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Section: Accepted Manuscriptmentioning

confidence: 99%

The secret life of conjugative relaxases

Guzmán-Herrador

Llosa

2019

Plasmid

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“…The TraI36 variant Y16F (TraI36(Y16F)) was crystallized with a bound ssDNA oligonucleotide (5#-TGGGGTGTĜ GTGCTTTTGGTGG-3#) corresponding to the complement of bases 127-148 of the F plasmid oriT sequence (Genbank accession number U01159 [Frost et al, 1994]). The sequence begins 8 bases 5# to nic (denoted by "^"), the site of specific ssDNA cleavage by TraI (Matson and Morton, 1991;Reygers et al, 1991). Throughout this paper, bases are numbered from the 5# end of the crystallization oligonucleotide, starting at number 1.…”

Section: Overall Conformation Of the Structurementioning

confidence: 99%

“…In vivo and in vitro experimentation has shown that relaxase interactions with oriT DNA are sequence specific (Fekete and Frost, 2000;Matson and Morton, 1991;Matson et al, 1993;Reygers et al, 1991;Stern and Schildbach, 2001; Thompson et al, 1984;Waters et al, 1991;Willetts and Maule, 1986). The binding specificity of F TraI36, the 36 kDa relaxase domain of the TraI protein from conjugative protein F factor, is exquisite.…”

Section: Introductionmentioning

confidence: 99%

Inter- and Intramolecular Determinants of the Specificity of Single-Stranded DNA Binding and Cleavage by the F Factor Relaxase

et al. 2005

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The TraI protein of conjugative plasmid F factor binds and cleaves a single-stranded region of the plasmid prior to transfer to a recipient. TraI36, an N-terminal TraI fragment, binds ssDNA with a subnanomolar K(D) and remarkable sequence specificity. The structure of the TraI36 Y16F variant bound to ssDNA reveals specificity determinants, including a ssDNA intramolecular 3 base interaction and two pockets within the protein's binding cleft that accommodate bases in a knob-into-hole fashion. Mutagenesis results underscore the intricate design of the binding site, with the greatest effects resulting from substitutions for residues that both contact ssDNA and stabilize protein structure. The active site architecture suggests that the bound divalent cation, which is essential for catalysis, both positions the DNA by liganding two oxygens of the scissile phosphate and increases the partial positive charge on the phosphorus to enhance nucleophilic attack.

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“…However, none of these proteins appear to require the replication protein A (RP-A) which has been shown to be involved in DNA replication and recombination in yeast and other eukaryotes (Wold & Kelly, 1987;Heyer & Kolodner, 1989;Heyer et al, 1990;Erdlieetal., 1991). As there are eight separate DNA helicases in E. coli, it is quite likely that there will be a number of DNA helicases in eukaryotic cells with unique functions and mechanisms of action (Lahue & Matson, 1988;Matson & Morton, 1991;Matson, 1991;Wood & Matson, 1987).…”

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confidence: 99%

Characterization of the DNA-dependent ATPase and a DNA unwinding activity associated with the yeast DNA polymerase .alpha. complex

Biswas

Ewing²,

Biswas³

1993

Biochemistry

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We have analyzed the ATPase and dATPase activities associated with the yeast DNA polymerase alpha complex. The ATPase/dATPase was primarily a single-stranded DNA-dependent ATPase. Analysis of the stimulatory effect of a large number of DNA substrates demonstrated that polynucleotides longer than 60 nucleotides (nts) had the maximal effect. The stimulation by oligonucleotides smaller than 60 nts, in general, decreased proportionally with decreased length of the oligomer. Poly- or oligopyrimidines were twice as stimulatory as the poly- or oligopurines of the same length. In addition to DNA, replication protein A (RP-A), a single-stranded DNA (ssDNA) binding protein, also stimulated the ATPase activity. Photo-cross-linking of the ATP binding component of the pol alpha complex to [alpha-32P]ATP at 0 degree C resulted in the exclusive labeling of a 90-kDa polypeptide. The labeling was inhibited by ATP and dATP but not by any other ribo- or deoxynucleotides, which suggest that the 90-kDa polypeptide is specific for ATP/dATP binding and possibly the active site for the ATPase/dATPase. We have also reported here a novel DNA unwinding activity associated with the multiprotein complex of DNA polymerase alpha. The complex was able to unwind M13mp19 ssDNA hybridized to an oligonucleotide (17-60 nucleotides long) with a protruding 3'-terminus. Regardless of the size of the duplex, the DNA unwinding was significantly stimulated by RP-A, while RP-A itself did not have any DNA unwinding activity. Consequently, it appeared that the DNA polymerase alpha complex possessed a putative RP-A-dependent helicase activity.(ABSTRACT TRUNCATED AT 250 WORDS)

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Escherichia coli DNA helicase I catalyzes a site- and strand-specific nicking reaction at the F plasmid oriT

Cited by 86 publications

References 29 publications

The secret life of conjugative relaxases

The secret life of conjugative relaxases

Inter- and Intramolecular Determinants of the Specificity of Single-Stranded DNA Binding and Cleavage by the F Factor Relaxase

Characterization of the DNA-dependent ATPase and a DNA unwinding activity associated with the yeast DNA polymerase .alpha. complex

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