Mechanical Constraints on Hin Subunit Rotation Imposed by the Fis/Enhancer System and DNA Supercoiling during Site-Specific Recombination

Dhar, Gautam; Heiss, John K.; Johnson, Reid C.

doi:10.1016/j.molcel.2009.05.020

Cited by 36 publications

(92 citation statements)

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“…In the invertasome, the recombinase dimers form a tetramer interacting with accessory DNA architectural protein FIS, whereas in the resolvosome the enzymatically active tetramer is associated with inactive recombinase tetramers and/or DNA architectural proteins bound at the adjacent accessory/regulatory DNA sites (Mouw et al 2008;Johnson 2015;Rice 2015). Consistent with the notion that DNA supercoiling increases the probability of juxtaposition of three DNA sites at branch points (Vologodskii and Cozzarelli 1996), the invertasome is assembled at a branch point in the negatively supercoiled DNA substrate on juxtaposition of three DNA sites, two recombination sites bound by the invertase tetramer, and the FIS-bound recombinational enhancer sequence (Johnson and Bruist 1989;Dhar et al 2009). This specificity of structural organization acts as a Btopological filter^imposing control on the alignment of recombination sites in the synaptic complex and, thus, in conjunction with stored negative superhelicity, imparting directionality on the reaction (Crisona et al 1994;Dhar et al 2009).…”

Section: H-ns Nucleoprotein Complexesmentioning

confidence: 53%

“…Consistent with the notion that DNA supercoiling increases the probability of juxtaposition of three DNA sites at branch points (Vologodskii and Cozzarelli 1996), the invertasome is assembled at a branch point in the negatively supercoiled DNA substrate on juxtaposition of three DNA sites, two recombination sites bound by the invertase tetramer, and the FIS-bound recombinational enhancer sequence (Johnson and Bruist 1989;Dhar et al 2009). This specificity of structural organization acts as a Btopological filter^imposing control on the alignment of recombination sites in the synaptic complex and, thus, in conjunction with stored negative superhelicity, imparting directionality on the reaction (Crisona et al 1994;Dhar et al 2009). DNA supercoiling energy plays an important role in this process, since any attempt to properly align recombination sites arranged in improper orientation in the substrate DNA (e.g., in the case of sites organized as direct repeats, instead of inverted repeats in an inversion substrate) would impose an energetic penalty by demanding introduction of an extra loop into the DNA.…”

Section: H-ns Nucleoprotein Complexesmentioning

confidence: 53%

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The regulatory role of DNA supercoiling in nucleoprotein complex assembly and genetic activity

Muskhelishvili

Travers

2016

Biophys Rev

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We argue that dynamic changes in DNA supercoiling in vivo determine both how DNA is packaged and how it is accessed for transcription and for other manipulations such as recombination. In both bacteria and eukaryotes, the principal generators of DNA superhelicity are DNA translocases, supplemented in bacteria by DNA gyrase. By generating gradients of superhelicity upstream and downstream of their site of activity, translocases enable the differential binding of proteins which preferentially interact with respectively more untwisted or more writhed DNA. Such preferences enable, in principle, the sequential binding of different classes of protein and so constitute an essential driver of chromatin organization.Keywords DNA supercoiling . DNA translocases . Chromatin organization . Superhelicity gradients . DNA untwisting . DNAwrithing Dynamic changes of DNA supercoiling act as a driving force behind the alterations of genetic activity and DNA compaction both in eukaryotes and prokaryotes. Both these processes involve formation of spatially organized nucleoprotein structures by DNA architectural proteins. Whereas in eukaryotes the paradigmal example is the histone octamer, in prokaryotes a similar function is attributed to a small class of highly abundant nucleoid-associated proteins. We argue that, depending on the primary sequence organization, the changes of superhelicity elicit various alterations of local DNA geometry, while these, in turn, facilitate the assembly of distinct nucleoprotein structures pertinent to genetic function. Genome-wide conversion of the superhelical energy into various three-dimensional DNA structures thus acts as an analogue code coordinating the energy supply with the genetic expression of the chromosome.Recent studies make it increasingly clear that the DNA double helix carries at least two types of encoded information and that these include the well-known genetic code and the structural information determining the form and physical properties of the double helix itself. Since both these information types are inscribed in the same DNA sequence, and yet one is discrete whereas the other is continuous, they have been respectively dubbed the digital and analog DNA codes (Marr et al. 2008;Travers et al. 2012;Muskhelishvili and Travers 2013). The potential of the DNA to adopt distinct configurations is strongly modulated by DNA supercoiling, which is increasingly recognized as one of the major forces coordinating the cellular DNA transactions in both prokaryotes (including Archaea) and eukaryotes.DNA supercoiling can be generated by the simple application of torque to the double helix (Strick et al. 1998) but, importantly, because the DNA molecule itself possesses chirality-it is, after all, a right-handed double helix-the local structures stabilized by changes in superhelical density depend on both the sign and strength of the applied torque. For example, both positive and negative torque (respectively overand underwinding of the double helix) can change the intrinsic coiling of ...

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Section: H-ns Nucleoprotein Complexesmentioning

confidence: 53%

Section: H-ns Nucleoprotein Complexesmentioning

confidence: 53%

The regulatory role of DNA supercoiling in nucleoprotein complex assembly and genetic activity

Muskhelishvili

Travers

2016

Biophys Rev

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“…In their natural context, small serine recombinases, such as the transposon resolvases and DNA invertases, stop after one round of strand exchange (1,25,26), even though the catalytic module (comprising two crossover sites synapsed by a recombinase tetramer) is fully competent for indefinite rounds of rotation (27). This selectivity is imposed by regulatory components involving accessory DNA sites and proteins (see introductory paragraphs).…”

Section: Discussionmentioning

confidence: 99%

Gated rotation mechanism of site-specific recombination by ϕC31 integrase

Olorunniji

Buck

Colloms

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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Integrases, such as that of the Streptomyces temperate bacteriophage ϕC31, promote site-specific recombination between DNA sequences in the bacteriophage and bacterial genomes to integrate or excise the phage DNA. ϕC31 integrase belongs to the serine recombinase family, a large group of structurally related enzymes with diverse biological functions. It has been proposed that serine integrases use a "subunit rotation" mechanism to exchange DNA strands after double-strand DNA cleavage at the two recombining att sites, and that many rounds of subunit rotation can occur before the strands are religated. We have analyzed the mechanism of ϕC31 integrase-mediated recombination in a topologically constrained experimental system using hybrid "phes" recombination sites, each of which comprises a ϕC31 att site positioned adjacent to a regulatory sequence recognized by Tn3 resolvase. The topologies of reaction products from circular substrates containing two phes sites support a right-handed subunit rotation mechanism for catalysis of both integrative and excisive recombination. Strand exchange usually terminates after a single round of 180°rotation. However, multiple processive "360°rotation" rounds of strand exchange can be observed, if the recombining sites have nonidentical base pairs at their centers. We propose that a regulatory "gating" mechanism normally blocks multiple rounds of strand exchange and triggers product release after a single round.T he large serine recombinase ϕC31 integrase (605 amino acids) promotes integration of the bacteriophage ϕC31 genome into the Streptomyces host chromosome by recombination between a phage site attP and a bacterial site attB, resulting in an integrated prophage flanked by two recombinant sites, attL and attR (1, 2). The prophage can remain dormant for many generations in this lysogenic state until it is excised by integrasemediated recombination between attL and attR, reforming attP and attB sites. The att sites (∼50 bp) each comprise sequences recognized by integrase flanking a central 2-bp overlap sequence at which crossover occurs (3-5) ( Fig. 1C and Fig. S1). Two integrase subunits bind to each att site, forming on-site dimers. Two att sites that are to recombine are then brought together by dimer-dimer interactions. ϕC31 integrase and related serine integrases catalyze efficient recombination between attP and attB, but not between the prophage sites attL and attR, or any other pairs of sites. However, in the presence of a phage-encoded recombination directionality factor, integrase specificity is altered; attL × attR recombination is efficient and attP × attB recombination is inhibited (4, 6, 7). There is considerable interest in this group of recombinases as tools for applications in biotechnology and genetic manipulation (8).Previous studies on the mechanism of recombination by small serine recombinases have led to a "subunit rotation" model for strand exchange (9-11). In this model, cleavage of all four DNA strands in a synaptic complex of the two recombining sites cre...

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“…But the recombination catalyzed by these “activated” enzymes has no directionality (either inversion or deletion) and may go through a different reaction pathway from wildtype recombinases (Arnold et al, 1999; Sarkis et al, 2001; Kamtekar et al, 2006; Gehman et al, 2008; Dhar et al, 2009). …”

Section: Site-specific Recombination By Yrs and Srsmentioning

confidence: 99%

Topoisomerases and site-specific recombinases: similarities in structure and mechanism

Yang

2010

Critical Reviews in Biochemistry and Molecular Biology

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The processes of DNA topoisomerization and site-specific recombination are fundamentally similar: DNA cleavage by forming a phospho-protein covalent linkage, DNA topological rearrangement, and DNA ligation coupled with protein regeneration. Type IB DNA topoisomerases are structurally and mechanistically homologous to tyrosine recombinases. Both enzymes nick DNA double helices independent of metal ions, form 3´-phosphotyrosine intermediates, and rearrange the free 5´ ends relative to the uncut strands by swiveling. In contrast, serine recombinases generate 5´-phospho-serine intermediates. A 180° relative rotation of the two halves of a 100kDa terameric serine recombinase and DNA complex has been proposed as the mechanism of strand exchange. Here I propose an alternative mechanism. Interestingly, the catalytic domain of serine recombinases has structural similarity to the TOPRIM domain, conserved among all Type IA and Type II topoisomerases and responsible for metal binding and DNA cleavage. TOPRIM topoisomerases also cleave DNA to generate 5´-phosphate and 3´-OH groups. Based on the existing biochemical data and crystal structures of Topoisomerase II and serine recombinases bound to pre- and post-cleavage DNA, I suggest a strand passage mechanism for DNA recombination by serine recombinases. This mechanism is reminiscent of DNA topoisomerization and does not require subunit rotation.

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Mechanical Constraints on Hin Subunit Rotation Imposed by the Fis/Enhancer System and DNA Supercoiling during Site-Specific Recombination

Cited by 36 publications

References 36 publications

The regulatory role of DNA supercoiling in nucleoprotein complex assembly and genetic activity

The regulatory role of DNA supercoiling in nucleoprotein complex assembly and genetic activity

Gated rotation mechanism of site-specific recombination by ϕC31 integrase

Topoisomerases and site-specific recombinases: similarities in structure and mechanism

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