Difference between revisions of "Part:BBa K3520005"
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<b><h1 style="color:green">cmcax: gene for Bacterial Cellulose upregulation</h1></b><br /><br />
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__NOTOC__
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<b>LONG DESCRIPTION</b><br /><br />
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<b>Project-General</b><br /><br />
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The current part is designed by iGEM Athens 2020 team during the project MORPHÆ. In this project, Flavobacteria were used to produce a non-cellular structurally coloured biomaterial which would require the secretion of a biomolecule that Flavobacteria do not normally secrete. Our hypothesis is that the formed matrix will have a structure similar to that of the biofilm and thus, it will provide the material with macroscopically the same colouration properties as the biofilm.
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<br /><br />
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<b>Operon related</b><br /><br />
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<partinfo>BBa_K3520005 short</partinfo>
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Our initial idea was to use bacterial cellulose, as an appropriate biomaterial, because of its unique properties, robustness, and biodegradability. The genes responsible for its production were selected from the bcs operon of <i>Komagataeibacter xylinus</i> (GenBank Acc. No. X54676.1), the most efficient bacterial cellulose producer, which consists of four genes, bcsA, bcsB, bcsC and bcsD. The bcsABCD operon encodes membrane-associated proteins that allow BC fibres to span through the membrane. Once the bcsABCD operon expression is triggered, BcsA and BcsB proteins form the BcsAB complex, which binds its substrate, UDP-glucose, at an intracellular glycosyltransferase (GT) domain and is the active core of cellulose synthase. This is followed by the secretion of BC fibres through pores and passageways formed by BcsC and BcsD proteins.Cmcax, CcpAx, cellulose synthase, BcsC, and BcsD are the biocatalysts of UDP-glucose transformation to cellulose. Two main applications of cellulose in biosciences are scaffolds for tissue engineering and generally in biomedicine.<br /><br />
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<br><br>
  
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<partinfo>BBa_K3520005 SequenceAndFeatures</partinfo>
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<br><br>
  
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<b>cmcax</b><br /><br />
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This part codes for the Cmcax protein, an assisting protein in cellulose biosynthesis in Bacteria. It is codon optimised for <i>Flavobacterium johnsoniae</i> UW101
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Cmcax was suggested to play a pivotal role in cellulose ribbon assembly as Cmcax is localized to the outer membrane surface and cell culture.
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<br/><br/>
  
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<b>SOURCE OF THIS PART</b><br /><br />
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=Description=
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The nucleotide sequences of the bacterial cellulose operon come from the strain <i>Komagataeibacter xylinus</i> and GenBank database (Acc.No.X54676.1). <i>K.xylinus</i> is a member of the acetic acid bacteria, a group of Gram-negative aerobic bacteria that produce acetic acid during fermentation. <br /><br />
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<br>
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It encodes an endo-b-1,4-glucanase, and thus exhibits cellulose-hydrolyzing activity. It has been shown that a small addition of active Cmcax increases bacterial cellulose synthesis. It may also exhert regulatory functions.
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Cmcax was suggested to play a pivotal role in cellulose ribbon assembly as Cmcax is localized to the outer membrane surface and cell culture. This CDS originates from a very productive cellulose synthesising bacterium, <i>Komagataeibacter xylinus</i> (GenBank Acc. No. X54676.1).
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<br><br>
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==The Bacterial Bcs Operon==
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<br>
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The putative operon consists of four genes, bcsA, bcsB, bcsC and bcsD. It encodes membrane-associated proteins that can catalyse extracellular Bacterial Cellulose synthesis in vivo. Once the bcsABCD operon expression is triggered, BcsA and BcsB proteins form the BcsAB complex, which binds its substrate, UDP-glucose, at an intracellular glycosyltransferase (GT) domain. This complex is the active core of the cellulose synthase. This is followed by the secretion of BC fibres through pores and passageways formed by BcsC and BcsD proteins. Co-expression with Cmcax and CcpAx have shown increased production. The cellulose synthase, BcsC, BcsD, Cmcax, and CcpAx are the biocatalysts of UDP-glucose transformation to cellulose. Two main applications of cellulose in biosciences are scaffolds for tissue engineering and generally in biomedicine.
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<br><br>
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=Athens 2020=
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<br>
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The current part is designed by iGEM Athens 2020 team during the project MORPHÆ. In this project, Flavobacteria were used to produce a non-cellular structurally coloured biomaterial which would require the secretion of a biomolecule that Flavobacteria do not normally secrete. Our hypothesis is that the formed matrix will have a structure similar to that of the biofilm and thus, it will provide the material with macroscopically the same colouration properties as the biofilm.
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 +
 
 +
=SOURCE OF THIS PART=
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<br>
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The nucleotide sequences of the bacterial cellulose operon come from the strain <i>Komagataeibacter xylinus</i> and GenBank database (Acc.No.X54676.1). <i>K.xylinus</i> is a member of the acetic acid bacteria, a group of Gram-negative aerobic bacteria that produce acetic acid during fermentation.
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<br><br>
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=Useful Links:=
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<br>
  
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<b>Useful Links:</b><br /><br />
 
 
NCBI taxonomy:<br /><br />
 
NCBI taxonomy:<br /><br />
 
https://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=28448&lvl=3&lin=f&keep=1&srchmode=1&unlock<br /><br />
 
https://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=28448&lvl=3&lin=f&keep=1&srchmode=1&unlock<br /><br />
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Codon optimization table:<br /><br />
 
Codon optimization table:<br /><br />
 
https://www.kazusa.or.jp/codon/cgi-bin/showcodon.cgi?species=376686&fbclid=IwAR0gwwrIarZsiYhWvHPc2BKy-iB_2OM-DPB5I2HYJZwBNiasmlLXWK87PwM<br /><br />
 
https://www.kazusa.or.jp/codon/cgi-bin/showcodon.cgi?species=376686&fbclid=IwAR0gwwrIarZsiYhWvHPc2BKy-iB_2OM-DPB5I2HYJZwBNiasmlLXWK87PwM<br /><br />
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<br><br>
  
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<b>REFERENCES</b><br /><br />
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=REFERENCES=
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<br>
  
 
Braun, T., Khubbar, M., Saffarini, D., & McBride, M. (2005). Flavobacterium johnsoniae Gliding Motility Genes Identified by mariner Mutagenesis. Journal Of Bacteriology, 187(20), 6943-6952. doi: 10.1128/jb.187.20.6943-6952.2005
 
Braun, T., Khubbar, M., Saffarini, D., & McBride, M. (2005). Flavobacterium johnsoniae Gliding Motility Genes Identified by mariner Mutagenesis. Journal Of Bacteriology, 187(20), 6943-6952. doi: 10.1128/jb.187.20.6943-6952.2005
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Römling, U., & Galperin, M. (2015). Bacterial cellulose biosynthesis: diversity of operons, subunits, products, and functions. Trends In Microbiology, 23(9), 545-557. doi: 10.1016/j.tim.2015.05.005
 
Römling, U., & Galperin, M. (2015). Bacterial cellulose biosynthesis: diversity of operons, subunits, products, and functions. Trends In Microbiology, 23(9), 545-557. doi: 10.1016/j.tim.2015.05.005
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Naoki Sunagawa, Takaaki Fujiwara, Takanori Yoda, Shin Kawano, Yasuharu Satoh, Min Yao,
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Kenji Tajima, and Tohru Dairi1, Cellulose complementing factor (Ccp) is a new member of the cellulose synthase
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complex (terminal complex) in Acetobacter xylinum, Journal of Bioscience and Bioengineering
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VOL. 115 No. 6, 607e612, 2013

Latest revision as of 03:07, 28 October 2020


cmcax-gene for Bacterial Cellulose upregulation


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]



This part codes for the Cmcax protein, an assisting protein in cellulose biosynthesis in Bacteria. It is codon optimised for Flavobacterium johnsoniae UW101



Description


It encodes an endo-b-1,4-glucanase, and thus exhibits cellulose-hydrolyzing activity. It has been shown that a small addition of active Cmcax increases bacterial cellulose synthesis. It may also exhert regulatory functions. Cmcax was suggested to play a pivotal role in cellulose ribbon assembly as Cmcax is localized to the outer membrane surface and cell culture. This CDS originates from a very productive cellulose synthesising bacterium, Komagataeibacter xylinus (GenBank Acc. No. X54676.1).

The Bacterial Bcs Operon


The putative operon consists of four genes, bcsA, bcsB, bcsC and bcsD. It encodes membrane-associated proteins that can catalyse extracellular Bacterial Cellulose synthesis in vivo. Once the bcsABCD operon expression is triggered, BcsA and BcsB proteins form the BcsAB complex, which binds its substrate, UDP-glucose, at an intracellular glycosyltransferase (GT) domain. This complex is the active core of the cellulose synthase. This is followed by the secretion of BC fibres through pores and passageways formed by BcsC and BcsD proteins. Co-expression with Cmcax and CcpAx have shown increased production. The cellulose synthase, BcsC, BcsD, Cmcax, and CcpAx are the biocatalysts of UDP-glucose transformation to cellulose. Two main applications of cellulose in biosciences are scaffolds for tissue engineering and generally in biomedicine.

Athens 2020


The current part is designed by iGEM Athens 2020 team during the project MORPHÆ. In this project, Flavobacteria were used to produce a non-cellular structurally coloured biomaterial which would require the secretion of a biomolecule that Flavobacteria do not normally secrete. Our hypothesis is that the formed matrix will have a structure similar to that of the biofilm and thus, it will provide the material with macroscopically the same colouration properties as the biofilm.


SOURCE OF THIS PART


The nucleotide sequences of the bacterial cellulose operon come from the strain Komagataeibacter xylinus and GenBank database (Acc.No.X54676.1). K.xylinus is a member of the acetic acid bacteria, a group of Gram-negative aerobic bacteria that produce acetic acid during fermentation.

Useful Links:


NCBI taxonomy:

https://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=28448&lvl=3&lin=f&keep=1&srchmode=1&unlock

GenBank link:

https://www.ncbi.nlm.nih.gov/nuccore/X54676.1

Codon optimisation bank:

http://genomes.urv.es/OPTIMIZER/?fbclid=IwAR0ALbP_C8UVY4itvYdNX8b5KYYUM5ulQojz8UJAK6Zj5llobNNxE-jYmXQ

Codon optimization table:

https://www.kazusa.or.jp/codon/cgi-bin/showcodon.cgi?species=376686&fbclid=IwAR0gwwrIarZsiYhWvHPc2BKy-iB_2OM-DPB5I2HYJZwBNiasmlLXWK87PwM



REFERENCES


Braun, T., Khubbar, M., Saffarini, D., & McBride, M. (2005). Flavobacterium johnsoniae Gliding Motility Genes Identified by mariner Mutagenesis. Journal Of Bacteriology, 187(20), 6943-6952. doi: 10.1128/jb.187.20.6943-6952.2005

Buldum, G., Bismarck, A., & Mantalaris, A. (2017). Recombinant biosynthesis of bacterial cellulose in genetically modified Escherichia coli. Bioprocess And Biosystems Engineering, 41(2), 265-279. doi: 10.1007/s00449-017-1864-1

Johansen, V., Catón, L., Hamidjaja, R., Oosterink, E., Wilts, B., & Rasmussen, T. et al. (2018). Genetic manipulation of structural color in bacterial colonies. Proceedings Of The National Academy Of Sciences, 115(11), 2652-2657. doi: 10.1073/pnas.1716214115

McBride, M., & Kempf, M. (1996). Development of techniques for the genetic manipulation of the gliding bacterium Cytophaga johnsonae. Journal Of Bacteriology, 178(3), 583-590. doi: 10.1128/jb.178.3.583-590.1996

Nakamura, Y. (2000). Codon usage tabulated from international DNA sequence databases: status for the year 2000. Nucleic Acids Research, 28(1), 292-292. doi: 10.1093/nar/28.1.292

Omadjela, O., Narahari, A., Strumillo, J., Melida, H., Mazur, O., Bulone, V., & Zimmer, J. (2013). BcsA and BcsB form the catalytically active core of bacterial cellulose synthase sufficient for in vitro cellulose synthesis. Proceedings Of The National Academy Of Sciences, 110(44), 17856-17861. doi: 10.1073/pnas.1314063110

Römling, U., & Galperin, M. (2015). Bacterial cellulose biosynthesis: diversity of operons, subunits, products, and functions. Trends In Microbiology, 23(9), 545-557. doi: 10.1016/j.tim.2015.05.005

Naoki Sunagawa, Takaaki Fujiwara, Takanori Yoda, Shin Kawano, Yasuharu Satoh, Min Yao, Kenji Tajima, and Tohru Dairi1, Cellulose complementing factor (Ccp) is a new member of the cellulose synthase complex (terminal complex) in Acetobacter xylinum, Journal of Bioscience and Bioengineering VOL. 115 No. 6, 607e612, 2013