Revision as of 11:42, 16 October 2016

ButCoat with strong RBS

Usage & Biology ( Description )

This composite part consists of RBS , our coding region ( ButCoaT ) and enables E.coli to secrete ButCoaT.

ButCoaT is an enzyme (Butyryl-CoA:acetate CoA-transferase) (EC 2.8.3.8) (Uniprot: C7GB37) which converts Acetyl-CoA to Butyrate and can be found in Roseburia intestinalis L1-82.

The equation of the reaction (acyl-CoA:acetate CoA-transferase) (KEGG Reaction No. : R01179) is given down below : Butanoyl-CoA + Acetate <=> Butanoic acid + Acetyl-CoA

RBS and our protein coding region (ButCoaT) are digested from our whole construct with the enzymes SpeI, Sall, Xbal.

For ButCoaT ;

E.Coli is synthesised suitable for Codon Bias. In the expression of pet28a (expression vector) , stop codon is removed in order to add HisTag.

Characterization

Design Notes

RBS and our protein coding region (ButCoaT) are digested from our whole construct with the enzymes SpeI, Sall, Xbal.

For ButCoaT ; E.Coli is synthesised suitable for Codon Bias. In the expression of pet28a (expression vector) , stop codon is removed in order to add HisTag.

Modeling

The figure above shows the increase in ButCoAT molecules over a time span of 10 seconds.

The figure above shows the molar increase in Butyrate concentration over a time span of 10 seconds.

Source

The ribosome binding site (B0034) that we’ve used is gotten from iGEM part library. Source of ButCoaT : Roseburia intestinalis L1-82

References

Hassig, C.A., Tong, J.K., Schreiber, S.L. (1997). Fiber-derived Butyrate and the Prevention of Colon Cancer

Roberto Berni Canani, Margherita Di Costanzo, Ludovica Leone, Monica Pedata, Rosaria Meli, Antonio Calignano. (2011). Potential beneficial affects of butyrate in intestinal and extraintestinal diseases

Duncan,S. H., Barcenilla, A., Stewart, C. S., Pryde, S. E., Flint, H. J. (2002). Acetate Utilization and Butyryl Coenzyme A (CoA):Acetate-CoA Transferase in ButyrateProducing Bacteria from the Human Large Intestine

Sequence : http://www.uniprot.org/uniprot/C7GB37?sort=score

Pathway : http://www.genome.jp/dbget-bin/www_bget?reaction+R00393+R01179+R01359+R07832

Picture : http://www.brenda-enzymes.org/structure.php?show=reaction&id=360609&type=S&displayType=marvin

RBS and our protein coding region (ButCoaT) are digested from our whole construct with the enzymes SpeI, Sall, Xbal.

For ButCoaT ;

E.Coli is synthesised suitable for Codon Bias. In the expression of pet28a (expression vector) , stop codon is removed in order to add HisTag.

Source

The ribosome binding site (B0034) that we’ve used is gotten from iGEM part library. Source of ButCoaT : Roseburia intestinalis L1-82

References

Hassig, C.A., Tong, J.K., Schreiber, S.L. (1997). Fiber-derived Butyrate and the Prevention of Colon Cancer

Roberto Berni Canani, Margherita Di Costanzo, Ludovica Leone, Monica Pedata, Rosaria Meli, Antonio Calignano. (2011). Potential beneficial affects of butyrate in intestinal and extraintestinal diseases

Duncan,S. H., Barcenilla, A., Stewart, C. S., Pryde, S. E., Flint, H. J. (2002). Acetate Utilization and Butyryl Coenzyme A (CoA):Acetate-CoA Transferase in ButyrateProducing Bacteria from the Human Large Intestine

Sequence : http://www.uniprot.org/uniprot/C7GB37?sort=score

Pathway : http://www.genome.jp/dbget-bin/www_bget?reaction+R00393+R01179+R01359+R07832

Picture : http://www.brenda-enzymes.org/structure.php?show=reaction&id=360609&type=S&displayType=marvin

Sequence and Features