Difference between revisions of "Part:BBa K343100"
 
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__NOTOC__
 
== Native ''flhDC'' master regulon ==
 
== Native ''flhDC'' master regulon ==
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This is only a coding sequence. The generator part is [https://parts.igem.org/Part:BBa_K343004 K343004] The native FlhDC operon has an internal Pst1 site positioned 822bp in the sequence. To use this part a silent mutation (T to C) will have to be introduced. <br>
 
=== Background ===
 
=== Background ===
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The flagella regulon in Escherichia coli is composed of at least 50 genes organized in no less than 14 ope-rons that all contribute to the synthesis and operation of flagella. The operons are synthesized in a three-level transcriptional cascade where the FlhDC operon is the master regulator at the top of the cascade. The flagella regulon is tightly controlled by nutritional and environmental conditions, E. coli starved of ami-no acids showed temporarily decrease of the flagella regulon transcripts which are needed for the synthesis and operation of the flagellum.[http://onlinelibrary.wiley.com/doi/10.1111/j.1365-2958.2009.06939.x/full (1)]
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The synthesis and assembly of flagella are regulated by the transcriptional cascade composed of three levels of gene products (class I, -II and –III). Class I genes consist of a single operon encoding the proteins FlhD and FlhC that form a multimeric (FlhD4C2) transcriptional activation complex. This ‘master regulator’ stimulates transcription by binding upstream of Class II promoters. Class II genes encode proteins that assemble to form the basal body and hook of the flagellum, as well as the fliA gene that encodes the alternative σ factor σ28, also called σF. σ28 binds to RNA polymerase (RNAP) core enzyme and directs it to Class III promoters. Class III genes encode the rest of the structural genes of the flagellum, including fliC encoding flagellin, as well as the chemotaxis apparatus. [http://onlinelibrary.wiley.com/doi/10.1111/j.1365-2958.2009.06939.x/full (1)]
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<br>
 
<br>
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It has been shown that overexpression of the FlhDC operon restores motility in mutants that have been made immotile [http://jb.asm.org/cgi/content/short/181/24/7500 (2)]. Also, overexpression of FlhDC in the E. coli K12 strain MG1655 made the cells hypermotile.[http://iai.asm.org/cgi/content/abstract/75/7/3315 (3)]
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More than 80% of all known bacterial species express flagella [https://parts.igem.org/Part:BBa_K343004#References [1]]. The synthesis of the flagella is a complex and energy consuming process, so the expression of the involved proteins is tightly regulated by the extracellular environment. One of the most well studied flagella synthesis systems is that of ''Eschericia coli''. Here the expression of the master regulator ''FlhDC'' operon is controlled i.e. by pH, salt concentration or the availability of nutrients. Apart from initiation of flagella synthesis FlhDC is also a repressor of cell metabolism as cell growth and flagellation does not occur at the same time. FlhDC is a hexameric transcription factor that consists of four FlhD subunits and two FlhC subunits (FlhD<sub>4</sub>C<sub>2</sub>). It constitutes the first class in the three classed flagella synthesis cascade. Class II consist of genes encoding proteins that make up the basal body, as well as the alternative transcription factor, σ<sup>28</sup>, which is responsible for the expression of the class III genes. These genes in turn encodes the proteins that composes the tail filaments.
 
<br><br>
 
<br><br>
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Studies of insertion elements (IS) upstream of the ''flhDC'' operon, has shown that an upregulation of the expression of the operon is responsible for the motility of the ''E. coli'' MG1655 strain. [https://parts.igem.org/Part:BBa_K343004#References [2]]It has also been shown that overexpression of the ''FlhDC'' operon restores motility in mutants that have been made immotile [https://parts.igem.org/Part:BBa_K343004#References [3]].
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<br>
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</p>
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==Refrences==
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<p style="text-align: justify;">
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<br>
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# O. Soutourina , PN. Bertin [http://www.ncbi.nlm.nih.gov/pubmed/14550943 Regulation cascade of flagellar expression in Gram-negative bacteria.]<br>
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# O. Soutourina, A. Kolb, E. Krin, C. Laurent-Winter, S. Rimsky, A. Danchin, and P. Bertin[http://jb.asm.org/cgi/content/short/181/24/7500 Multiple Control of Flagellum Biosynthesis in Escherichia coli: Role of H-NS Protein and the Cyclic AMP-Catabolite Activator Protein Complex in Transcription of the flhDC Master Operon]<br>
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# Eric J. Gauger, Mary P. Leatham, Regino Mercado-Lubo, David C. Laux, Tyrrell Conway, and Paul S. Cohen [http://iai.asm.org/cgi/content/abstract/75/7/3315 Role of Motility and the flhDC Operon in Escherichia coli MG1655 Colonization of the Mouse Intestine{triangledown}]<br>
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</p>
 
<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here
 
===Usage and Biology===
 
===Usage and Biology===
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<span class='h3bb'>Sequence and Features</span>
 
<span class='h3bb'>Sequence and Features</span>
 
<partinfo>BBa_K343100 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K343100 SequenceAndFeatures</partinfo>
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=== Safety ===
 
=== Safety ===
  

Latest revision as of 23:09, 27 October 2010

Native flhDC master regulon

This is only a coding sequence. The generator part is K343004 The native FlhDC operon has an internal Pst1 site positioned 822bp in the sequence. To use this part a silent mutation (T to C) will have to be introduced.

Background


More than 80% of all known bacterial species express flagella [1]. The synthesis of the flagella is a complex and energy consuming process, so the expression of the involved proteins is tightly regulated by the extracellular environment. One of the most well studied flagella synthesis systems is that of Eschericia coli. Here the expression of the master regulator FlhDC operon is controlled i.e. by pH, salt concentration or the availability of nutrients. Apart from initiation of flagella synthesis FlhDC is also a repressor of cell metabolism as cell growth and flagellation does not occur at the same time. FlhDC is a hexameric transcription factor that consists of four FlhD subunits and two FlhC subunits (FlhD4C2). It constitutes the first class in the three classed flagella synthesis cascade. Class II consist of genes encoding proteins that make up the basal body, as well as the alternative transcription factor, σ28, which is responsible for the expression of the class III genes. These genes in turn encodes the proteins that composes the tail filaments.

Studies of insertion elements (IS) upstream of the flhDC operon, has shown that an upregulation of the expression of the operon is responsible for the motility of the E. coli MG1655 strain. [2]It has also been shown that overexpression of the FlhDC operon restores motility in mutants that have been made immotile [3].

Refrences


  1. O. Soutourina , PN. Bertin [http://www.ncbi.nlm.nih.gov/pubmed/14550943 Regulation cascade of flagellar expression in Gram-negative bacteria.]
  2. O. Soutourina, A. Kolb, E. Krin, C. Laurent-Winter, S. Rimsky, A. Danchin, and P. Bertin[http://jb.asm.org/cgi/content/short/181/24/7500 Multiple Control of Flagellum Biosynthesis in Escherichia coli: Role of H-NS Protein and the Cyclic AMP-Catabolite Activator Protein Complex in Transcription of the flhDC Master Operon]
  3. Eric J. Gauger, Mary P. Leatham, Regino Mercado-Lubo, David C. Laux, Tyrrell Conway, and Paul S. Cohen [http://iai.asm.org/cgi/content/abstract/75/7/3315 Role of Motility and the flhDC Operon in Escherichia coli MG1655 Colonization of the Mouse Intestine{triangledown}]

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal PstI site found at 821
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal PstI site found at 821
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal PstI site found at 821
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal PstI site found at 821
    Illegal AgeI site found at 609
  • 1000
    COMPATIBLE WITH RFC[1000]

Safety

[http://iai.asm.org/cgi/reprint/75/7/3315 Role of Motility and the flhDC Operon in Escherichia coli MG1655 Colonization of the Mouse Intestine]

[http://www.ncbi.nlm.nih.gov/pmc/articles/PMC93725/ The Yersinia enterocolitica Motility Master Regulatory Operon, flhDC, Is Required for Flagellin Production, Swimming Motility, and Swarming Motility]

[http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0008029 Repression of Invasion Genes and Decreased Invasion in a High-Level Fluoroquinolone-Resistant Salmonella Typhimurium Mutant]