User:Christina Manner/Sandbox 810: Difference between revisions

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Quorum sensing is used by bacteria to regulate gene expression depending on cell-population density <ref name = Miller> Miller M.B., Bassler C. L. (2001): ''Quorum Sensing in Bacteria.'' In: ''Annu. Rev. Microbiol.'' 55; 165-199 </ref>. Therefore, Bacteria use small hormone-like proteins called autoinducers <ref name = Waters> Waters C. M., Bassler C. L. (2005): ''Quorum Sensing: Cell-to-Cell Communication in Bacteria.'' In: ''Annu. Rev. Cell Biol.'' 21; 319-346 </ref>. These autoinducers increase in concentration in connection to increasing cell density <ref name = Miller />. Reaching a minimal threshold stimulatory concentration, the autoinducers activate gene regulation processes <ref name = Miller />. This quorum sensing becomes beneficial as soon as it is performed by many cells <ref name = Waters />. Quorum sensing is used by Gram-negative as well as Gram-positive bacteria and occurs within and between bacterial species <ref name = Miller />. The communication via quorum sensing may have been a first step of multi-cellularity and makes the distinction between eukaryotes and prokaryotes more complex <ref name = Miller /> <ref name = Waters />.  

TraR is member of the quorum-sensing transcription factor family called LuxR including the Helix-Turn-Helix motif <ref name = Volpari> A.,Volpari C., Di Marco S. (2004): ''Crystal Structure of the Quorum-sensing Protein TraM and Its Interaction with the Transcriptional Regulator TraR.'' In: ''J. Biol. Chem.''  279(23); 24291–96 </ref>, that is present in many DNA binding proteins, e.g. Cro, CAP or the λ-repressor <ref name = Brennan> Brennan RG., Matthews BW. (1989): The Helix-Turn-Helix DNA Binding Motif. In: J. of Biol. Chem. 264(4); 1903-06 </ref>. In presence of its autoinducer AAI, TraR regulates genes connected to the tumor inducing (Ti) plasmid  <ref name = Chai />. When a certain cell density of Agrobacterium tumefaciens is reached, the transfer of the Ti-plasmid is induced <ref name = Volpari/>. There are two proteins that influence this transfer: TraR and TraI. The TraI gene encodes AAI. The absence of AAI causes rapid proteolysis of TraR  <ref name = Chai /> , which implies, that AAI protects TraR from degradation <ref name = Vannini />. TraR itself activates the Ti-plasmid tra genes [7].  

In general TraR works as a <scene name='57/574296/Dimer/2'>homo-dimer</scene>, but the monomers are differently elongated which leads to a general asymmetric structure of the dimer. Each TraR monomer consists of 234 amino acids. In this <scene name='57/574296/1h0m/2'>structure</scene>, <scene name='57/574296/1h0m/3'>two TraR dimers</scene> binding the tra box are shown. A and C are in the same way elongated, as well as B and D. Each monomer possesses its own <scene name='57/574296/Ligand_binding_site/2'>ligand binding domain</scene> as well as a <scene name='57/574296/Dna_binding_domain/1'>DNA-binding domain</scene>. Thus, all in all four autoinducer molecules are bound to the TraR proteins at one tra box.

The dimers AB and CD interact with each other via molecular interactions between B and C. Additionally, the palindromic sequence of the tra box causes a base stacking between the beginning and the end of the sequence.  

As A and B are slightly different, there is an dimeric asymmetry that has two consequences for the function. At first, the <scene name='57/574296/N-ter_dimer/3'>N-terminal parts</scene> of the two monomers have different positions. The N-terminal part of A is between the ligand-binding domain and the DNA-binding domain in the center of the protein. In opposition to that, the N-terminal part of B is located externally. Moreover, there is a <scene name='57/574296/Neg_charged_dna_binding_region/2'>distribution of charge</scene> at the surface of the dimer. Because of that, the DNA-binding domain forms a long and basic region for the interaction with DNA, whereas the C-terminal residues form a positively charged patch exposed to the solvent. This region might be involved in protein-protein interaction with TraM <ref name = Volpari/>. TraM binds to TraR and prevents it from binding to the DNA and therefore prevents the activation of the transcription of the tra genes <ref name = Volpari/>.

==='''Ligand binding domain'''===

Following the ligand binding domain, the residues 163 to 175 form the <scene name='57/574296/Linker/3'>linker</scene> to the DNA binding domain. The residues 166 to 169 are disordered and cannot be seen in the structural model.  

==='''DNA binding domain'''===

The DNA binding domain contains four <scene name='57/574296/Dna_binding_domain_alpha/1'> α-helices </scene> (α7,  α8, α9,  α10) and is provided by the C-terminal part of the protein by the residues 176 to 234. In monomer A or C, the first residues of α7, the last residues of α8 and the residues 232 to 234 form polar bonds with the N-terminal part of the ligand binding domain. In the B or D monomer, no such interactions were observed. Besides, helices α8 and α9 form the <scene name='57/574296/Hth/1'>HTH</scene> (Helix-Turn-Helix) motif.  

TraR is the activator for the conjugal transfer of Ti-plamids. [8] In nature, Agrobacterium tumefaciens uses the transfer of the Ti-plasmid to manipulate dicotyledonous plants into producing metabolites as nutrients for the bacteria. [7] TraR activates the tra genes if AAI is present and also activates the transcription of traR and traI. [8] The tra genes are responsible for the conjugal transfer.  

In biotechnology, the whole transfer system of the Ti-plasmid can be used to transfer DNA into dicotelydonous plant cells and to integrate that DNA into the plant genome. Naturally, Agrobacterium tumefaciens only infects dicotelydonous plants and the systems to perform this are well established. But there were also some experiments that proved a transformation using the ti-plasmid in monocotyledons. [8]</div>