Difference between revisions of "Part:BBa K2259010"
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__TOC__
 
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[[Image:Scheme of rop.jpeg|right|250px|thumb|<b>Figure 1. </b>Structure of the ColE1 Rop protein, at 1.7 angstroms resolution.<ref>Banner DW, Kokkinidis M, Tsernoglou D. Structure of the ColE1 Rop protein at 1.7 Å resolution. J Mol Biol. 1987 m.;196(3):657–75.</ref>]]
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[[Image:Scheme of rop.jpeg|right|225px|thumb|<b>Figure 1. </b> Main principles of ColE1 plasmid family replication. Rop protein interaction region marked in red square.]]
  
 
=Introduction=
 
=Introduction=
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<b>Repressor of primer (ROP)</b> is a small dimeric protein that participates in ColE1 plasmid family copy number control, by increasing affinity between two complementary RNAs - RNA I (Replication inhibitor) and RNA II (Replication activator). <ref>Castagnoli L, Scarpa M, Kokkinidis M, Banner DW, Tsernoglou D, Cesareni G. Genetic and structural analysis of the ColE1 Rop (Rom) protein. The EMBO Journal. 1989;8(2):621-629.</ref> By increasing affinity of the two RNA molecules Rop decreases the rate of of plasmid replication initiation events.
 
<b>Repressor of primer (ROP)</b> is a small dimeric protein that participates in ColE1 plasmid family copy number control, by increasing affinity between two complementary RNAs - RNA I (Replication inhibitor) and RNA II (Replication activator). <ref>Castagnoli L, Scarpa M, Kokkinidis M, Banner DW, Tsernoglou D, Cesareni G. Genetic and structural analysis of the ColE1 Rop (Rom) protein. The EMBO Journal. 1989;8(2):621-629.</ref> By increasing affinity of the two RNA molecules Rop decreases the rate of of plasmid replication initiation events.
  
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[[Image:Rop protein 3d small.gif|right|500px|frame|<b>Figure 1. </b>Structure of the ColE1 Rop protein, at 1.7 angstroms resolution.<ref>Banner DW, Kokkinidis M, Tsernoglou D. Structure of the ColE1 Rop protein at 1.7 Å resolution. J Mol Biol. 1987 m.;196(3):657–75.</ref>]]
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[[Image:Rop protein 3d small.gif|right|500px|frame|<b>Figure 2. </b>Structure of the ColE1 Rop protein, at 1.7 angstroms resolution.<ref>Banner DW, Kokkinidis M, Tsernoglou D. Structure of the ColE1 Rop protein at 1.7 Å resolution. J Mol Biol. 1987 m.;196(3):657–75.</ref>]]
  
 
Rop dimer is a bundle of four tightly packed alpha helices that are held by hydrophobic interactions (Fig. 1).
 
Rop dimer is a bundle of four tightly packed alpha helices that are held by hydrophobic interactions (Fig. 1).
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===Rop protein in SynORI===
 
===Rop protein in SynORI===
 
Rop protein does not recognise specific sequences of RNA I and RNA II molecules, but instead recognises the RNA I - RNA II kissing loop complex secondary structures. That means it can act as a <b>global copy number modulator</b>, which bypasses the selective control of each plasmid group.
 
Rop protein does not recognise specific sequences of RNA I and RNA II molecules, but instead recognises the RNA I - RNA II kissing loop complex secondary structures. That means it can act as a <b>global copy number modulator</b>, which bypasses the selective control of each plasmid group.
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  <b>For example:</b> You have a ''two-plasmid system'', with specific RNA I concentrations set so that first plasmid group has an average  
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  <b>For example:</b> You have a ''two-plasmid system'', with specific RNA I concentrations set so that
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copy number of ''100'', and another group at ''50'' copies. Rop protein can be used to <b>globally lower the copy number of each group </b> -  
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first plasmid group has an average copy number of ''100'', and another group at ''50'' copies. Rop  
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from 100 to 50 and from 50 to 25 copies respectively.  
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protein can be used to <b>globally lower the copy number of each group </b> - from 100 to 50 and  
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The degree of copy number reduction depends on Rop concentration in a cell.
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from 50 to 25 copies respectively. The degree of copy number reduction depends
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on Rop concentration in a cell.
  
 
=Characterization of Rop protein (Vilnius-Lithuania 2017)=
 
=Characterization of Rop protein (Vilnius-Lithuania 2017)=

Revision as of 00:45, 19 October 2017


Rop protein - global copy number inhibitor (SynORI framework)

Rop (also known as repressor of primer) is a small protein responsible for keeping the copy number of ColE1 and related bacterial plasmids low in E. coli. Inhibits plasmid replication of all SynORI framework plasmid groups non-specifically, also other ColE1 replicon plasmids.



Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Figure 1. Main principles of ColE1 plasmid family replication. Rop protein interaction region marked in red square.

Introduction

Biology

Repressor of primer (ROP) is a small dimeric protein that participates in ColE1 plasmid family copy number control, by increasing affinity between two complementary RNAs - RNA I (Replication inhibitor) and RNA II (Replication activator). [1] By increasing affinity of the two RNA molecules Rop decreases the rate of of plasmid replication initiation events.

Figure 2. Structure of the ColE1 Rop protein, at 1.7 angstroms resolution.[2]

Rop dimer is a bundle of four tightly packed alpha helices that are held by hydrophobic interactions (Fig. 1).

Usage with SynORI (Framework for multi-plasmid systems)

About SynORI

SynORI is a framework for multi-plasmid systems created by Vilnius-Lithuania 2017 which enables quick and easy workflow with multiple plasmids, while also allowing to freely pick and modulate copy number for every unique plasmid group! Read more about [http://2017.igem.org/Team:Vilnius-Lithuania SynORI here]!

Rop protein in SynORI

Rop protein does not recognise specific sequences of RNA I and RNA II molecules, but instead recognises the RNA I - RNA II kissing loop complex secondary structures. That means it can act as a global copy number modulator, which bypasses the selective control of each plasmid group. 

For example: You have a two-plasmid system, with specific RNA I concentrations set so that
first plasmid group has an average copy number of 100, and another group at 50 copies. Rop 
protein can be used to globally lower the copy number of each group  - from 100 to 50 and 
from 50 to 25 copies respectively. The degree of copy number reduction depends
on Rop concentration in a cell.

Characterization of Rop protein (Vilnius-Lithuania 2017)

References

  1. ↑ Castagnoli L, Scarpa M, Kokkinidis M, Banner DW, Tsernoglou D, Cesareni G. Genetic and structural analysis of the ColE1 Rop (Rom) protein. The EMBO Journal. 1989;8(2):621-629.
  2. ↑ Banner DW, Kokkinidis M, Tsernoglou D. Structure of the ColE1 Rop protein at 1.7 Å resolution. J Mol Biol. 1987 m.;196(3):657–75.