User:Chloe Paul/Sandbox 1: Difference between revisions

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== Introduction ==

Replication Terminator Protein (RTP) from Bacillus Subtilis is a protein of current scientific investigation in terms of its symmetric and asymmetric nature, its ability to bind DNA and the mechanism upon which it terminates DNA replication. Belonging to a group of Replication Terminator Proteins that are commonly found in prokaryotes (in particular within the Bacillaceae family) (~~Pfam~~); RTP is often compared to another protein with a similar intracellular function, Termination Utilisation Sequence (Tus) from E. coli. RTP has been shown to exist in both symmetric (in solution and when bound to palindromic DNA sequences) and asymmetric states (when bound to native DNA). Its structural properties have proven to be integral to its function; as it must be able to bind DNA and have polarity (despite it being a homomeric dimer) in order to specifically block DNA replication from one direction and yet be “permissive” at the other end.

Replication Terminator Protein (RTP) from Bacillus Subtilis is a protein of current scientific investigation in terms of its symmetric and asymmetric nature, its ability to bind DNA and the mechanism upon which it terminates DNA replication. Belonging to a group of Replication Terminator Proteins that are commonly found in prokaryotes (in particular within the Bacillaceae family)<ref> R.D. Finn, J. Mistry, J. Tate, P. Coggill, A. Heger, J.E. Pollington, O.L. Gavin, P. Gunesekaran, G. Ceric, K. Forslund, L. Holm, E.L. Sonnhammer, S.R. Eddy, A. Bateman The Pfam protein families database [http://pfam.sanger.ac.uk/family/PF02334#tabview=tab0] Nucleic Acids Research (2010) Database Issue 38:D211-222</ref>; RTP is often compared to another protein with a similar intracellular function, Termination Utilisation Sequence (Tus) from E. coli. RTP has been shown to exist in both symmetric (in solution and when bound to palindromic DNA sequences) and asymmetric states (when bound to native DNA). Its structural properties have proven to be integral to its function; as it must be able to bind DNA and have polarity (despite it being a homomeric dimer) in order to specifically block DNA replication from one direction and yet be “permissive” at the other end.

== The Structure of RTP ==

RTP has been found to exist as a symmetric protein in solution as a homomeric dimer through crystal structure determination ~~(Bussiere, 1995)~~. These identical subunits/monomers each contain four α helices (α1, α2, α3, α4) and three β strands (β1, β2, β3) along with a disordered N-terminal region. When the two subunits come together and the two α4 helices align, they form the dimer with an overall rectangle shape of 66Å x 35 Å x 30 Å. The long C-terminal helices (α4) are involved three interactions: contributing to the hydrophobic core (residues 93-103), an antiparallel coiled-coil structure (the two α4 helices coming together) and the contributing to the hydrophobic core of the other monomer (residue 122). Both monomers still remain structurally similar when they form the dimer in solution. It should be noted that when in solution the flexible loop between β2 and β3 are able to assume different ~~conformations~~. ~~(Bussiere, 1995)~~

RTP has been found to exist as a symmetric protein in solution as a homomeric dimer through crystal structure determination <ref name="bussiere">PMID: 7867072</ref>.These identical subunits/monomers each contain four α helices (α1, α2, α3, α4) and three β strands (β1, β2, β3) along with a disordered N-terminal region. When the two subunits come together and the two α4 helices align, they form the dimer with an overall rectangle shape of 66Å x 35 Å x 30 Å <ref name="bussiere" />. The long C-terminal helices (α4) are involved three interactions: contributing to the hydrophobic core (residues 93-103), an antiparallel coiled-coil structure (the two α4 helices coming together) and the contributing to the hydrophobic core of the other monomer (residue 122)<ref name="bussiere" />. Both monomers still remain structurally similar when they form the dimer in solution. It should be noted that when in solution the flexible loop between β2 and β3 are able to assume different conformation <ref name="bussiere" />.

== RTP binding to DNA ==

RTP has been of interest in terms of its specific binding to DNA because it doesn’t use the common DNA structural motifs such as a basic leucine zipper, zinc finger or helix-turn-helix motif. It has been established that RTP like Tus, is sequence specific, as it binds as the Ter sites (comprising of two sequences that are imperfect inverted repeats (Vivian et al, 2007 ~~and Bussiere 1995)~~). This means that RTP needs to be able to recognise specific bases in the helical DNA structure by reading the exposed edges of the bases located in the major and minor grooves of DNA not involved in pairing. Structurally RTP interacts with DNA through its alpha helices in the major grooves, its anti-parallel β-strands in the minor grooves and the flexible N-terminal regions wrapping with non-specific ionic interactions around the DNA (Wilce et al, 2001).

RTP has been of interest in terms of its specific binding to DNA because it doesn’t use the common DNA structural motifs such as a basic leucine zipper, zinc finger or helix-turn-helix motif. It has been established that RTP like Tus, is sequence specific, as it binds as the Ter sites (comprising of two sequences that are imperfect inverted repeats (Vivian et al, 2007 <ref name="bussiere" />). This means that RTP needs to be able to recognise specific bases in the helical DNA structure by reading the exposed edges of the bases located in the major and minor grooves of DNA not involved in pairing. Structurally RTP interacts with DNA through its alpha helices in the major grooves, its anti-parallel β-strands in the minor grooves and the flexible N-terminal regions wrapping with non-specific ionic interactions around the DNA (Wilce et al, 2001).

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==The Fork Arrest Mechanism==

As previously noted the role of RTP is to terminate replication of the bacterial chromosome. It was originally assumed that the role of RTP was simply to arrest the replication fork allowing the DNA to cleanly separate (Wake 1997.) The proposed mechanism noted that the replication fork is only able to disrupt the RTP/ter interaction when approaching the A-site. This explains the polarity of the mechanism. However recent research has indicated a more complex mechanism involving interactions between the bound RTP and the replication fork helicase. The results of this research have confirmed a RTP/DnaB interaction in vivo, further suggesting this interaction plays an important role in replication fork arrest (Gautam 2001.) This has lead to the development of a new helicase-specific model involving protein-protein interactions between the replication fork helicase and RTP which arrests the replication fork when it approaches from the appropriate direction (Kaplan 2009.)

==References==

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 == Introduction ==
-Replication Terminator Protein (RTP) from Bacillus Subtilis is a protein of current scientific investigation in terms of its symmetric and asymmetric nature, its ability to bind DNA and the mechanism upon which it terminates DNA replication. Belonging to a group of Replication Terminator Proteins that are commonly found in prokaryotes (in particular within the Bacillaceae family) (Pfam); RTP is often compared to another protein with a similar intracellular function, Termination Utilisation Sequence (Tus) from E. coli. RTP has been shown to exist in both symmetric (in solution and when bound to palindromic DNA sequences) and asymmetric states (when bound to native DNA). Its structural properties have proven to be integral to its function; as it must be able to bind DNA and have polarity (despite it being a homomeric dimer) in order to specifically block DNA replication from one direction and yet be “permissive” at the other end.
+Replication Terminator Protein (RTP) from Bacillus Subtilis is a protein of current scientific investigation in terms of its symmetric and asymmetric nature, its ability to bind DNA and the mechanism upon which it terminates DNA replication. Belonging to a group of Replication Terminator Proteins that are commonly found in prokaryotes (in particular within the Bacillaceae family)<ref> R.D. Finn, J. Mistry, J. Tate, P. Coggill, A. Heger, J.E. Pollington, O.L. Gavin, P. Gunesekaran, G. Ceric, K. Forslund, L. Holm, E.L. Sonnhammer, S.R. Eddy, A. Bateman The Pfam protein families database [http://pfam.sanger.ac.uk/family/PF02334#tabview=tab0] Nucleic Acids Research (2010)  Database Issue 38:D211-222</ref>; RTP is often compared to another protein with a similar intracellular function, Termination Utilisation Sequence (Tus) from E. coli. RTP has been shown to exist in both symmetric (in solution and when bound to palindromic DNA sequences) and asymmetric states (when bound to native DNA). Its structural properties have proven to be integral to its function; as it must be able to bind DNA and have polarity (despite it being a homomeric dimer) in order to specifically block DNA replication from one direction and yet be “permissive” at the other end.
 == The Structure of RTP ==
-RTP has been found to exist as a symmetric protein in solution as a homomeric dimer through crystal structure determination (Bussiere, 1995). These identical subunits/monomers each contain four α helices (α1, α2, α3, α4) and three β strands (β1, β2, β3) along with a disordered N-terminal region. When the two subunits come together and the two α4 helices align, they form the dimer with an overall rectangle shape of 66Å x 35 Å x 30 Å. The long C-terminal helices (α4) are involved three interactions: contributing to the hydrophobic core (residues 93-103), an antiparallel coiled-coil structure (the two α4 helices coming together) and the contributing to the hydrophobic core of the other monomer (residue 122). Both monomers still remain structurally similar when they form the dimer in solution. It should be noted that when in solution the flexible loop between β2 and β3 are able to assume different conformations. (Bussiere, 1995)
+RTP has been found to exist as a symmetric protein in solution as a homomeric dimer through crystal structure determination <ref name="bussiere">PMID: 7867072</ref>.These identical subunits/monomers each contain four α helices (α1, α2, α3, α4) and three β strands (β1, β2, β3) along with a disordered N-terminal region. When the two subunits come together and the two α4 helices align, they form the dimer with an overall rectangle shape of 66Å x 35 Å x 30 Å <ref name="bussiere" />. The long C-terminal helices (α4) are involved three interactions: contributing to the hydrophobic core (residues 93-103), an antiparallel coiled-coil structure (the two α4 helices coming together) and the contributing to the hydrophobic core of the other monomer (residue 122)<ref name="bussiere" />. Both monomers still remain structurally similar when they form the dimer in solution. It should be noted that when in solution the flexible loop between β2 and β3 are able to assume different conformation <ref name="bussiere" />.
 == RTP binding to DNA ==
-RTP has been of interest in terms of its specific binding to DNA because it doesn’t use the common DNA structural motifs such as a basic leucine zipper, zinc finger or helix-turn-helix motif. It has been established that RTP like Tus, is sequence specific, as it binds as the Ter sites (comprising of two sequences that are imperfect inverted repeats (Vivian et al, 2007 and Bussiere 1995)). This means that RTP needs to be able to recognise specific bases in the helical DNA structure by reading the exposed edges of the bases located in the major and minor grooves of DNA not involved in pairing. Structurally RTP interacts with DNA through its alpha helices in the major grooves, its anti-parallel β-strands in the minor grooves and the flexible N-terminal regions wrapping with non-specific ionic interactions around the DNA (Wilce et al, 2001).
+RTP has been of interest in terms of its specific binding to DNA because it doesn’t use the common DNA structural motifs such as a basic leucine zipper, zinc finger or helix-turn-helix motif. It has been established that RTP like Tus, is sequence specific, as it binds as the Ter sites (comprising of two sequences that are imperfect inverted repeats (Vivian et al, 2007 <ref name="bussiere" />). This means that RTP needs to be able to recognise specific bases in the helical DNA structure by reading the exposed edges of the bases located in the major and minor grooves of DNA not involved in pairing. Structurally RTP interacts with DNA through its alpha helices in the major grooves, its anti-parallel β-strands in the minor grooves and the flexible N-terminal regions wrapping with non-specific ionic interactions around the DNA (Wilce et al, 2001).
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 ==The Fork Arrest Mechanism==
 As previously noted the role of RTP is to terminate replication of the bacterial chromosome. It was originally assumed that the role of RTP was simply to arrest the replication fork allowing the DNA to cleanly separate (Wake 1997.) The proposed mechanism noted that the replication fork is only able to disrupt the RTP/ter interaction when approaching the A-site. This explains the polarity of the mechanism. However recent research has indicated a more complex mechanism involving interactions between the bound RTP and the replication fork helicase. The results of this research have confirmed a RTP/DnaB interaction in vivo, further suggesting this interaction plays an important role in replication fork arrest (Gautam 2001.) This has lead to the development of a new helicase-specific model involving protein-protein interactions between the replication fork helicase and RTP which arrests the replication fork when it approaches from the appropriate direction (Kaplan 2009.)
+==References==
+<references />