Limits...
The carboxy-terminal αN helix of the archaeal XerA tyrosine recombinase is a molecular switch to control site-specific recombination.

Serre MC, El Arnaout T, Brooks MA, Durand D, Lisboa J, Lazar N, Raynal B, van Tilbeurgh H, Quevillon-Cheruel S - PLoS ONE (2013)

Bottom Line: Surprisingly, XerA C-terminal αN helices dock in cis in a groove that, in bacterial tyrosine recombinases, accommodates in trans αN helices of neighbour monomers in the Holliday junction intermediates.Deletion of the XerA C-terminal αN helix does not impair cleavage of suicide substrates but prevents recombination catalysis.We propose that the enzymatic cycle of XerA involves the switch of the αN helix from cis to trans packing, leading to (i) repositioning of the catalytic Tyr in the active site in cis and (ii) dimer stabilisation via αN contacts in trans between monomers.

View Article: PubMed Central - PubMed

Affiliation: Institut de Génétique et Microbiologie, Université Paris-Sud, Orsay, France. marie-claude.serre@igmors.u-psud.fr

ABSTRACT
Tyrosine recombinases are conserved in the three kingdoms of life. Here we present the first crystal structure of a full-length archaeal tyrosine recombinase, XerA from Pyrococcus abyssi, at 3.0 Å resolution. In the absence of DNA substrate XerA crystallizes as a dimer where each monomer displays a tertiary structure similar to that of DNA-bound Tyr-recombinases. Active sites are assembled in the absence of dif except for the catalytic Tyr, which is extruded and located equidistant from each active site within the dimer. Using XerA active site mutants we demonstrate that XerA follows the classical cis-cleavage reaction, suggesting rearrangements of the C-terminal domain upon DNA binding. Surprisingly, XerA C-terminal αN helices dock in cis in a groove that, in bacterial tyrosine recombinases, accommodates in trans αN helices of neighbour monomers in the Holliday junction intermediates. Deletion of the XerA C-terminal αN helix does not impair cleavage of suicide substrates but prevents recombination catalysis. We propose that the enzymatic cycle of XerA involves the switch of the αN helix from cis to trans packing, leading to (i) repositioning of the catalytic Tyr in the active site in cis and (ii) dimer stabilisation via αN contacts in trans between monomers.

Show MeSH

Related in: MedlinePlus

Cis or trans interactions of αN helices in Tyr-recombinases.The C-terminal domains of XerA and Cre are compared. αMN helices are in orange and catalytic Tyr are in red sticks. For Cre, M2 and N2 helices come from the neighbouring monomer. The groove occupied in trans by αN2 helix in Cre is the same as the groove occupied in cis by the αN helix of the same subunit in XerA.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3646895&req=5

pone-0063010-g007: Cis or trans interactions of αN helices in Tyr-recombinases.The C-terminal domains of XerA and Cre are compared. αMN helices are in orange and catalytic Tyr are in red sticks. For Cre, M2 and N2 helices come from the neighbouring monomer. The groove occupied in trans by αN2 helix in Cre is the same as the groove occupied in cis by the αN helix of the same subunit in XerA.

Mentions: The XerA structure revealed that the extreme C-terminal αN helix packs in cis into a groove located at the surface of the C-terminal domain (Figures 2 and 7). Comparison with the Cre C-terminal domain structure indicates that the same groove accommodates the αN helix delivered in trans from the neighbouring subunit (Figure 7). Notably the contact regions between αN helices and the core domain are similar in cis and trans packing (Figure 7). Within the Cre dimer bound to DNA, the αN helix of the first molecule (N1) is extruded from the catalytic domain, whereas its αM helix (M1) is oriented toward the catalytic pocket in cis (Figure 7). This conformation positions the catalytic Tyr into the catalytic site of the same monomer. Meanwhile, the αN helix of the second subunit (N2) tightly contacts in trans the first subunit. The non reciprocal swapping of αN helices contributes to the stabilisation of the dimer. The αN helix is also critical for contacts between XerC and XerD. In fact, interactions between the two recombinases are not critical for the initial steps of recombination but proved to be essential to recombine dif sites [16]. Within the XerA apo-dimer, positioning of the αN helix in cis prevents contacts between the αM helix and the catalytic site resulting in the exclusion of the catalytic Tyr from the active site (Figure 7). It is thus tempting to hypothesize that upon DNA binding, the two last helices (αMN) of the XerA C-terminal domain could relocate in trans and induce repositioning of the catalytic Tyr within the active site in cis.


The carboxy-terminal αN helix of the archaeal XerA tyrosine recombinase is a molecular switch to control site-specific recombination.

Serre MC, El Arnaout T, Brooks MA, Durand D, Lisboa J, Lazar N, Raynal B, van Tilbeurgh H, Quevillon-Cheruel S - PLoS ONE (2013)

Cis or trans interactions of αN helices in Tyr-recombinases.The C-terminal domains of XerA and Cre are compared. αMN helices are in orange and catalytic Tyr are in red sticks. For Cre, M2 and N2 helices come from the neighbouring monomer. The groove occupied in trans by αN2 helix in Cre is the same as the groove occupied in cis by the αN helix of the same subunit in XerA.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3646895&req=5

pone-0063010-g007: Cis or trans interactions of αN helices in Tyr-recombinases.The C-terminal domains of XerA and Cre are compared. αMN helices are in orange and catalytic Tyr are in red sticks. For Cre, M2 and N2 helices come from the neighbouring monomer. The groove occupied in trans by αN2 helix in Cre is the same as the groove occupied in cis by the αN helix of the same subunit in XerA.
Mentions: The XerA structure revealed that the extreme C-terminal αN helix packs in cis into a groove located at the surface of the C-terminal domain (Figures 2 and 7). Comparison with the Cre C-terminal domain structure indicates that the same groove accommodates the αN helix delivered in trans from the neighbouring subunit (Figure 7). Notably the contact regions between αN helices and the core domain are similar in cis and trans packing (Figure 7). Within the Cre dimer bound to DNA, the αN helix of the first molecule (N1) is extruded from the catalytic domain, whereas its αM helix (M1) is oriented toward the catalytic pocket in cis (Figure 7). This conformation positions the catalytic Tyr into the catalytic site of the same monomer. Meanwhile, the αN helix of the second subunit (N2) tightly contacts in trans the first subunit. The non reciprocal swapping of αN helices contributes to the stabilisation of the dimer. The αN helix is also critical for contacts between XerC and XerD. In fact, interactions between the two recombinases are not critical for the initial steps of recombination but proved to be essential to recombine dif sites [16]. Within the XerA apo-dimer, positioning of the αN helix in cis prevents contacts between the αM helix and the catalytic site resulting in the exclusion of the catalytic Tyr from the active site (Figure 7). It is thus tempting to hypothesize that upon DNA binding, the two last helices (αMN) of the XerA C-terminal domain could relocate in trans and induce repositioning of the catalytic Tyr within the active site in cis.

Bottom Line: Surprisingly, XerA C-terminal αN helices dock in cis in a groove that, in bacterial tyrosine recombinases, accommodates in trans αN helices of neighbour monomers in the Holliday junction intermediates.Deletion of the XerA C-terminal αN helix does not impair cleavage of suicide substrates but prevents recombination catalysis.We propose that the enzymatic cycle of XerA involves the switch of the αN helix from cis to trans packing, leading to (i) repositioning of the catalytic Tyr in the active site in cis and (ii) dimer stabilisation via αN contacts in trans between monomers.

View Article: PubMed Central - PubMed

Affiliation: Institut de Génétique et Microbiologie, Université Paris-Sud, Orsay, France. marie-claude.serre@igmors.u-psud.fr

ABSTRACT
Tyrosine recombinases are conserved in the three kingdoms of life. Here we present the first crystal structure of a full-length archaeal tyrosine recombinase, XerA from Pyrococcus abyssi, at 3.0 Å resolution. In the absence of DNA substrate XerA crystallizes as a dimer where each monomer displays a tertiary structure similar to that of DNA-bound Tyr-recombinases. Active sites are assembled in the absence of dif except for the catalytic Tyr, which is extruded and located equidistant from each active site within the dimer. Using XerA active site mutants we demonstrate that XerA follows the classical cis-cleavage reaction, suggesting rearrangements of the C-terminal domain upon DNA binding. Surprisingly, XerA C-terminal αN helices dock in cis in a groove that, in bacterial tyrosine recombinases, accommodates in trans αN helices of neighbour monomers in the Holliday junction intermediates. Deletion of the XerA C-terminal αN helix does not impair cleavage of suicide substrates but prevents recombination catalysis. We propose that the enzymatic cycle of XerA involves the switch of the αN helix from cis to trans packing, leading to (i) repositioning of the catalytic Tyr in the active site in cis and (ii) dimer stabilisation via αN contacts in trans between monomers.

Show MeSH
Related in: MedlinePlus