Difference between revisions of "Part:BBa K2052018"
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==Source==
 
==Source==
  
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Roseburia intestinalis L1-82
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We obtained the enzyme ButCoAT from Roseburia intestinalis L1-82 by doing codon bias to our E.coli.
  
  

Revision as of 09:30, 17 October 2016


Butyryl-CoA:acetate CoA-transferase

ButCoAT is an enzyme (Butyryl-CoA:acetate CoA-transferase) (EC 2.8.3.8) (Uniprot:C7GB37) which converts Acetyl-CoA (Acetyl-CoA is readily present in the cytosol and it can be directly converted into Butyrate) into Butyrate and can be found in Roseburia intestinalis L1-82. The equation of the reaction (acyl-CoA:acetate CoA-transferase) is given down below :

Pathwayson.png

Figure 1: Butanoyl-CoA + Acetate <=> Butanoic acid + Acetyl-CoA

ButCoaTStructure.gif

Figure 2: Crystal Structures of Acetobacter aceti Succinyl (Butyryl)-CoA:Acetate CoA-Transferase Reveal Specificity Determinants and Illustrate the Mechanism Used by Class I CoA-Transferases.(Mullins, E.A. et al., 2012)

Butyrate

Butyrate is a short chain fatty acid which is formed by bacterial fermentation of carbohydrates (Sengupta et al., 2005).

Butyrate putatively suppresses colorectal cancer (CRC). It induces apoptosis and differentiation, inhibits proliferation of tumorous cells in colon flora. At the intestinal level, butyrate plays an administrative part on the transepithelial liquid transport, improves mucosal aggravation and oxidative status, strengthens the epithelial protection obstruction, and regulates instinctive affectability and intestinal motility.

Modeling

caption

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

caption

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

Design Notes

E.coli is synthesized suitable for Codon Bias.

In the expression of pet28a (expression vector) , stop codon is removed for the addition of HisTag.

DNA Gel Analysis

Characterization

PCR Conformation of ButCoAT + pSB1C3

ButCoaTGelImage.jpeg

We have loaded the uncut version of K2052015 (RBS + ButCoAT) next to EcoRI single digested one. In the third lane, RBS ligated ButCoAT (K2052018) was loaded with its single digested version. Digested ones gave sharp lane at 3600 bp as we expected.

Source

We obtained the enzyme ButCoAT from Roseburia intestinalis L1-82 by doing codon bias to our E.coli.


Sequence and Features


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]


References

Weinstock G., Sodergren E., Clifton S., Fulton L., Fulton B., Courtney L., Fronick C., Harrison M., Strong C., Farmer C., Delahaunty K., Markovic C., Hall O., Minx P., Tomlinson C., Mitreva M., Nelson J., Hou S. Wilson R.K. (2009 , August ). UniProtKB - C7GB37 (C7GB37_9FIRM). Retrieved from http://www.uniprot.org/uniprot/C7GB37?sort=score

Sengupta, S., Muir, G.J., Gibson, P. R. (2005). Does Butyrate Protect from Colorectal Cancer?

Figure 1 ; BRENDA. Reaction catalyzed by acetate CoA-transferase (2.8.3.8). Retrieved from http://www.brenda-enzymes.org/structure.php?show=reaction&id=360609&type=S&displayType=marvin

Figure 2 ; SWISS-MODEL. Succinyl-CoA:acetate CoA-transferase (AarCH6-S71A) in complex with CoA. Retrieved from https://swissmodel.expasy.org/templates/4eu8.1