Part:BBa_K1462390:Design
pTDH3+CA+PDZ-ligand+tADH1
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 525
Illegal BglII site found at 1551 - 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
This recombinant consists of four parts, the constitutive promoter TDH3, the CA, the ligand of the scaffold PDZ and the teminator ADH1.
Carbonic anhydrases (or carbonate dehydratases) form a family of enzymes that catalyze the rapid interconversion of carbon dioxide and water to bicarbonate and protons (or vice versa), a reversible reaction that occurs relatively slowly in the absence of a catalyst. The active site of most carbonic anhydrases contains a zincion; they are therefore classified as metalloenzymes. The reaction catalyzed by carbonic anhydrase is: H2CO3→CO2 + H2O The reaction rate of carbonic anhydrase is one of the fastest of all enzymes, and its rate is typically limited by the diffusion rate of its substrates. The epsilon(ε) class of CAs occurs exclusively in bacteria in a few chemolithotrophs and marine cyanobacteria that contain cso-carboxysomes.
Source
The CA we use is a component of the carboxysome shell of Halothiobacillus neapolitanus c2. We got pTDH3 and tADH1 from kits, CA from a synthesis company and PDZ-ligand by PCR(PDZ-ligand sequence included in the reverse primer used to amplify CA).
References
Walter Bonaccia, Poh K. Tengb, Bruno Afonsoa, et al. : Modularity of a carbon-fixing protein organelle. PNAS 2012 vol. 109 , no. 2: 478-483.