Team:Groningen/Project/Promoters

From 2009.igem.org

(Difference between revisions)
(Introduction)
(Copper Induced Promoters)
Line 11: Line 11:
===Copper Induced Promoters===
===Copper Induced Promoters===
 +
 +
Promoter cusCp is associated with the two component system CusR and CusS for the copper induced transcription of genes involved in copper efflux (cusC, cusF, cusB and cusA, which is present on the genome of Escherichia coli str. K-12 substrain MG1655). The sequence shows the typical -10 and -35 region of the promoter and can be found through the following [http://biocyc.org/ECOLI/NEW-IMAGE?type=OPERON&object=TU0-1821 link]. A second region, located at -53.5 from the transcription start site, is thought to bind CusR. Upon binding of CusR, the RNA polymerase is able to recognize the site and attach itself, and can also be found in the same [http://biocyc.org/ECOLI/NEW-IMAGE?type=OPERON&object=TU0-1821 link].
 +
 +
*CusS, a sensory histidine kinase in a two-component regulatory system with CusR, is able to recognize copper ions, phosphorilate, and form a complex with CusR. It's a 480 amino acid long protein of which the sequence (aa and nt) can be found [http://www.genome.jp/dbget-bin/www_bget?eco+b0570 here] allong with other information.
 +
 +
*CusR, "Cu-sensing regulator", regulates genes related to the copper and silver efflux systems under '''anaerobic growth''' and under '''extreme copper stress''' in aerobic growth . It's a 227 amino acid long protein of which the sequence (aa and nt) can be found [http://www.genome.jp/dbget-bin/www_bget?eco+b0571 here] along with other information.
 +
 +
 +
Cu --> CusS --> +P --> CusR --> Activation of transription
 +
 +
 +
The toxicity of copper is largely due to its tendency to alternate between its cuprous, Cu(I), and cupric, Cu(II), oxidation states, differentiating copper from other trace metals, such as zinc or nickel. Under aerobic conditions, this redox cycling leads to the generation of highly reactive hydroxyl radicals that readily and efficiently damage biomolecules, such as DNA, proteins, and lipids.(needs a ref.)
The toxicity of copper is largely due to its tendency to alternate between its cuprous, Cu(I), and cupric, Cu(II), oxidation states, differentiating copper from other trace metals, such as zinc or nickel. Under aerobic conditions, this redox cycling leads to the generation of highly reactive hydroxyl radicals that readily and efficiently damage biomolecules, such as DNA, proteins, and lipids.(needs a ref.)
Line 24: Line 36:
*Ettema TJ,Brinkman AB,Lamers PP,Kornet NG,de Vos WM,van der Oost J. Molecular characterization of a conserved archaeal copper resistance (cop) gene cluster and its copper-responsive regulator in Sulfolobus solfataricus P2.Microbiology 2006 Jul;152(Pt 7):1969-79.
*Ettema TJ,Brinkman AB,Lamers PP,Kornet NG,de Vos WM,van der Oost J. Molecular characterization of a conserved archaeal copper resistance (cop) gene cluster and its copper-responsive regulator in Sulfolobus solfataricus P2.Microbiology 2006 Jul;152(Pt 7):1969-79.
Abstract: That CopT binds to the copMA promoter at multiple sites, both upstream and downstream of the predicted TATA-BRE site. Copper was found to specifically modulate the affinity of DNA binding by CopT. This study describes a copper-responsive operon in archaea, a new family of archaeal DNA-binding proteins, and supports the idea that this domain plays a prominent role in the archaeal copper response. A model is proposed for copper-responsive transcriptional regulation of the copMA gene cluster.
Abstract: That CopT binds to the copMA promoter at multiple sites, both upstream and downstream of the predicted TATA-BRE site. Copper was found to specifically modulate the affinity of DNA binding by CopT. This study describes a copper-responsive operon in archaea, a new family of archaeal DNA-binding proteins, and supports the idea that this domain plays a prominent role in the archaeal copper response. A model is proposed for copper-responsive transcriptional regulation of the copMA gene cluster.
-
 
===Arsenicum Induced Promoters===
===Arsenicum Induced Promoters===

Revision as of 10:06, 15 June 2009

Igemhomelogo.png

Introduction

A promoter is a part of DNA involved in the regulation of gene transcription by ribosomes. In general ribosomes tend to bind weakly to a strand of DNA until a suitable promoter is encountered and the binding becomes strong.

Different Promoters

In order to find different promoters to induce the different genes here is a list of databases for promoters:

  1. Regtransbase

Copper Induced Promoters

Promoter cusCp is associated with the two component system CusR and CusS for the copper induced transcription of genes involved in copper efflux (cusC, cusF, cusB and cusA, which is present on the genome of Escherichia coli str. K-12 substrain MG1655). The sequence shows the typical -10 and -35 region of the promoter and can be found through the following link. A second region, located at -53.5 from the transcription start site, is thought to bind CusR. Upon binding of CusR, the RNA polymerase is able to recognize the site and attach itself, and can also be found in the same link.

  • CusS, a sensory histidine kinase in a two-component regulatory system with CusR, is able to recognize copper ions, phosphorilate, and form a complex with CusR. It's a 480 amino acid long protein of which the sequence (aa and nt) can be found here allong with other information.
  • CusR, "Cu-sensing regulator", regulates genes related to the copper and silver efflux systems under anaerobic growth and under extreme copper stress in aerobic growth . It's a 227 amino acid long protein of which the sequence (aa and nt) can be found here along with other information.


Cu --> CusS --> +P --> CusR --> Activation of transription



The toxicity of copper is largely due to its tendency to alternate between its cuprous, Cu(I), and cupric, Cu(II), oxidation states, differentiating copper from other trace metals, such as zinc or nickel. Under aerobic conditions, this redox cycling leads to the generation of highly reactive hydroxyl radicals that readily and efficiently damage biomolecules, such as DNA, proteins, and lipids.(needs a ref.)

  • Mills SD,Lim CK,Cooksey DA. Purification and characterization of CopR, a transcriptional activator protein that binds to a conserved domain (cop box) in copper-inducible promoters of Pseudomonas syringae. Mol. Gen. Genet. 1994 Aug;244(4):341-51.

Abstract: The copper resistance (cop) operon promoter (Pcop) of Pseudomonas syringae is copper-inducible, and requires the regulatory genes copR and copS. Primer extension analysis identified the transcriptional initiation site of Pcop 59 bp 5' to the translational start site of copA.

  • Khunajakr N,Liu CQ,Charoenchai P,Dunn NW. A plasmid-encoded two-component regulatory system involved in copper-inducible transcription in Lactococcus lactis. Gene 1999 Mar;229(1-2):229-35.

Abstract: Two regulatory genes (lcoR and lcoS) were identified from a plasmid-borne lactococcal copper resistance determinant and characterized by transcriptional fusion to the promoterless chloramphenicol acetyltransferase gene (cat). The transcription start site involved in copper induction was mapped by primer extension.

  • Ettema TJ,Brinkman AB,Lamers PP,Kornet NG,de Vos WM,van der Oost J. Molecular characterization of a conserved archaeal copper resistance (cop) gene cluster and its copper-responsive regulator in Sulfolobus solfataricus P2.Microbiology 2006 Jul;152(Pt 7):1969-79.

Abstract: That CopT binds to the copMA promoter at multiple sites, both upstream and downstream of the predicted TATA-BRE site. Copper was found to specifically modulate the affinity of DNA binding by CopT. This study describes a copper-responsive operon in archaea, a new family of archaeal DNA-binding proteins, and supports the idea that this domain plays a prominent role in the archaeal copper response. A model is proposed for copper-responsive transcriptional regulation of the copMA gene cluster.

Arsenicum Induced Promoters

Because of the similarity to phosphate, sometimes arsenate is mistaken for phosphate, which is how it is introduced into living organisms, including E. coli, by the phosphate uptake system. Other molecules such as As(III) can also be introduced into the cells by various membrane transporters. (needs a ref.)

Requirements

  • Need for specificity towards the wanted metals
  • Usable in E.coli

Literature

  1. Nawapan S., Charoenlap N., Charoenwuttitam A., Saenkham P., Mongkolsuk S., and Vattanaviboon P., Functional and expression analyses of the cop operon required for copper resistance in Agrobacterium tumefaciens, 2009, J. Bacteriol. doi:10.1128/JB.00384-09