Replication Termination Protein: Difference between revisions

From Proteopedia
Jump to navigation Jump to search
← Older editNewer edit →
Michal Harel (talk | contribs)
31,761 edits
No edit summary
Michal Harel (talk | contribs)
31,761 edits
No edit summary
Line 1: Line 1:
[[Image:RTP.png|450px|right|thumb| Diagram of RTP monomer with secondary structure highlighted.]]
[[Image:RTP.png|450px|right|thumb| Diagram of RTP monomer with secondary structure highlighted.]]
The '''replication termination protein''' (RTP) is one of only two well-defined proteins known to be involved in arresting DNA replication forks, the other being a protein known as tus (termination utilisation substance) from E. coli <ref> Kamada K, Horiuchi T, Ohsumi K, Shimamoto N, Morkikawa K, (1996) Structure of a replication-terminator protein complexed with DNA.  Nature, 383:598-603 </ref>. RTP was discovered in Bacillus subtilis  and has been identified as a DNA binding protein of the winged helix family that forms a dimer of 29kDa. This dimeric form has been shown to have an exceptionally high affinity for its cognate binding sites( Kd ~10-11M-1)<ref> Wilce et. al. (2001) Structure of the RTP−DNA complex and the mechanism of polar replication fork arrest. Nature Structural Biology, 8:206-210 </ref>, otherwise known as Termination sites (Ter sites). These Ter sites are found in multiple locations in the B. subtilis genome <ref> Gautam A. et.al. (2001) A single domain of the replication termination protein of Bacillus subtilis is involved in arresting both DnaB helicase and RNA polymerase. Journal of Biological Chemistry, 276:23471-23479</ref>.  For more details see:<br />
The '''replication termination protein''' (RTP) is one of only two well-defined proteins known to be involved in arresting DNA replication forks, the other being a protein known as tus (termination utilisation substance) from E. coli <ref> Kamada K, Horiuchi T, Ohsumi K, Shimamoto N, Morkikawa K, (1996) Structure of a replication-terminator protein complexed with DNA.  Nature, 383:598-603 </ref>. RTP was discovered in ''Bacillus subtilis'' and has been identified as a DNA binding protein of the winged helix family that forms a dimer of 29kDa. This dimeric form has been shown to have an exceptionally high affinity for its cognate binding sites( Kd ~10-11M-1)<ref> Wilce et. al. (2001) Structure of the RTP−DNA complex and the mechanism of polar replication fork arrest. Nature Structural Biology, 8:206-210 </ref>, otherwise known as Termination sites (Ter sites). These Ter sites are found in multiple locations in the B. subtilis genome <ref> Gautam A. et.al. (2001) A single domain of the replication termination protein of Bacillus subtilis is involved in arresting both DnaB helicase and RNA polymerase. Journal of Biological Chemistry, 276:23471-23479</ref>.  For more details see:<br />
*[[RTP and Tus]]<br />
*[[RTP and Tus]]<br />
*[[Rtp and Tus DNA Binding]]<br />
*[[Rtp and Tus DNA Binding]]<br />

Revision as of 10:41, 4 August 2016

Diagram of RTP monomer with secondary structure highlighted.

The replication termination protein (RTP) is one of only two well-defined proteins known to be involved in arresting DNA replication forks, the other being a protein known as tus (termination utilisation substance) from E. coli [1]. RTP was discovered in Bacillus subtilis and has been identified as a DNA binding protein of the winged helix family that forms a dimer of 29kDa. This dimeric form has been shown to have an exceptionally high affinity for its cognate binding sites( Kd ~10-11M-1)[2], otherwise known as Termination sites (Ter sites). These Ter sites are found in multiple locations in the B. subtilis genome [3]. For more details see:


Termination of ReplicationTermination of Replication

B. subtilis genome showing origin of replication, polar Ter sites and both replication forks.

Bacterial replication like that found in Bacillus subtilis consists of two replication forks that travel in opposite directions around the same circular strand of DNA. These replication forks begin at the origin of replication (OriC) and travel in clockwise and anticlockwise directions [4]. To prevent the strand from being over replicated each strand needs to be terminated roughly opposite the OriC and then joined to form the complete duplicated strand of DNA. As such the Ter/RTP complexes have found to be polar in their action; that is that different Ter sites are capable of only blacking DNA replication from one direction only. Since Ter sites are found facing both forks of replication, and the combination of both are capable of stopping both strands of replication, they are often termed as replication traps. The arresting of replication is achieved by the binding of two dimers to each Ter site, with the two RTP binding sites referred to as site A and site B. Site B shows a higher affinity for RTP and binds first, with site A then being filled with another RTP dimer cooperatively. This differential affinity for RTP is cited as a possible reason for the observed polarity of the termination [5].

RTP StructureRTP Structure

RTP is a DNA binding protein from Bacillus Subtilis that uses a helix-loop-helix binding motif. In solution it shows a symmetric structure typical of the winged helix loop helix family, with an unstructured end, first alpha helix , unstructured loop that is equivalent to the first beta sheet , helix loop helix structure (-), 2 beta sheets with a connecting loop that makes up the 'wing' structure and an additional long alpha helix involved in dimerisation .[6]

Binding of Rtp to the assymetric B portion of the Ter site changes its conformation into an assymetric 'wing-up wing-down' structure.

The contacts upstream with rtp dimer bound to A-site while the : Contacts with phosphate backbone of downstream DNA

Assymetric RTP dimer bound to the B site in a Ter region (PDB code 2dpd)

Drag the structure with the mouse to rotate

Residues that bind in both monomers: Red

Residues that bind only in wing-up: Green

Residues that bind only in wing-down: Blue

Both monomers Unit Binding
Lys14 Wing-down Phosphate(13)
Gln15 Both Phosphate(14)
Arg16 Both Phosphate(13)
Tyr33 Wing-down Phosphate(3)
Leu35 both Phosphate(4)
Lys36 Wing-up Sugar(3)
Asn53 Both Phosphate(14)
His54 Both Guanine(5)
Thr55 Both Thymine(15)
Glu56 Wing-up Phosphate(13)
Tyr58 Both Phosphate(4/5)
Arg59 Both Guanine(14)
His62 Both Phosphate(5)
Gln72 Both Phosphate(5)
Lys77 Wing-down Phosphate(3)
Gln83 Wing-down Sugar(3)
Val86 Wind-down Phosphate(4)

[7]

Replication MechanismReplication Mechanism

Debate over the mechanism of Replication termination by RTP has led to a number of mechanisms being proposed. Original work on this question was hampered by an incorrect structure of RTP being released. This structure was produced in solution and was shown to be a symmetric dimer[8] Later crystallisation of the protein bound to DNA showed the RTP formed a asymmetric dimer when bound to cognate sequences [9] . One early study proposed that the asymmetric interaction between terminator protein and terminator DNA contributed to the observed polarity, with later studies showing that the RTP/Ter complex contained partially unwound DNA, suggesting a locked complex was responsible. More recent data has implicated that protein -protein interactions between RTP and the helicase is primarily responsible for termination and that this mechanism is modulated by the asymmetrical interactions described above [10].

3D structures of replication terminator protein3D structures of replication terminator protein

1bm9 – BsRTP – Bacillus subtilis
1j0r, 2dqr - BsRTP (mutant)
2dpd - BsRTP + DNA
1f4k, 2dpu, 2efw – BsRTP (mutant) + DNA
1ecr – Tus + DNA – Escherichia coli

ReferencesReferences

  1. ↑ Kamada K, Horiuchi T, Ohsumi K, Shimamoto N, Morkikawa K, (1996) Structure of a replication-terminator protein complexed with DNA. Nature, 383:598-603
  2. ↑ Wilce et. al. (2001) Structure of the RTP−DNA complex and the mechanism of polar replication fork arrest. Nature Structural Biology, 8:206-210
  3. ↑ Gautam A. et.al. (2001) A single domain of the replication termination protein of Bacillus subtilis is involved in arresting both DnaB helicase and RNA polymerase. Journal of Biological Chemistry, 276:23471-23479
  4. ↑ Noirot P (2007). "Replication of the Bacillus subtilis chromosome". In Graumann P. Bacillus: Cellular and Molecular Biology. Caister Academic Press. ISBN 978-1-904455-12-7
  5. ↑ Duggin I.G. (2006) DNA Replication Fork Arrest by the Bacillus subtilis RTP–DNA Complex Involves a Mechanism that Is Independent of the Affinity of RTP–DNA Binding. Journal of Molecular Biology, 361:1-6
  6. ↑ Vivian JP, Porter CJ, Wilce JA, Wilce MCJ, (2007) An asymmetric structure of the Bacillus subtilise Replication Terminator Protein in Complex with DNA. J. Mol. Bio, 370:481-491
  7. ↑ Vivian JP, Porter CJ, Wilce JA, Wilce MCJ, (2007) An asymmetric structure of the Bacillus subtilise Replication Terminator Protein in Complex with DNA. J. Mol. Bio, 370:481-491
  8. ↑ Bastia D. (1995) Crystal structure of the replication terminator protein from b. subtilis at 2.6 A. Cell 80: 651-660
  9. ↑ Wilce et. al. (2001) Structure of the RTP−DNA complex and the mechanism of polar replication fork arrest. Nature Structural Biology, 8:206-210
  10. ↑ Kaplan D.L., Bastia D. (2009). Mechanisms of polar arrest of a replication fork. Molecular Microbiology 72: 279-284

Proteopedia Page Contributors and Editors (what is this?)Proteopedia Page Contributors and Editors (what is this?)

Craig Mooney, Michal Harel, Alexander Berchansky, Joel L. Sussman
Retrieved from "https://proteopedia.openfox.io/wiki/index.php?title=Replication_Termination_Protein&oldid=2624705"