Difference between revisions of "Part:BBa K4863000"

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α-carbonic anhydrase from Helicobacter pylori 26695 (hpCA)

A-carbonic anhydrases are metalloenzymes responsible for catalyzing the interconversion of carbon dioxide (CO2) to bicarbonate ions (HCO3−). hpCA is an α-carbonic anhydrase isolated from the bacterium Helicobacter pylori 26695, codon optimized for expression in E. Coli BL21(DE3) (Zhu et al., 2022).

In our work, CA is expressed in E. Coli and purified for display on the surface of Synechocystis PCC6803 for converting carbon dioxide into bicarbonate ions, which then forms calcium carbonate (CaCO3) precipitates, thus achieving biomineralization and the production of biological concrete.

Usage and Biology

Carbonic anhydrases (CAs) can catalyze the biochemical process necessary for the biotechnological method of microbially induced calcite precipitation (MICP), which has gained increasing attention due to its potential in creating calcium carbonate precipitation for increasing the compressive strength of concrete. CAs catalyze the conversion of carbon dioxide to bicarbonate ions, and the ions can react with CaCl to form calcium carbonate deposition, which acts as an adhesive for increasing and maintaining compressive strength of concrete. hpCA, the α-carbonic anhydrase isolated from Helicobacter pylori strain 26695, has relatively high performance among the CAs characterized so far for expression in E. Coli. Additionally, it has been shown that surface display is necessary for maintaining the stability of the enzyme (Zhu et al., 2022).

A-CAs are naturally present in mammals and many Eubacteria species and is responsible for various biochemical processes including and not limiting to maintaining concentration of carbon dioxide or bicarbonate ions for carboxylation reactions in autotrophs, transporting carbon dioxide or bicarbonate ions in mammals, and regulating intracellular pH(Bury-Moné, 2008). HpCA is encoded by the open reading frame HP1186 in Helicobacter pylori 26695. It is attached to the cell surface and is responsible for creating a CO2-rich environment which is necessary for survival of the organism(Chirica, 2002).

Source

Helicobacter pylori 26695

Characterization

   <figure>
       <img src="" width="500">
       <figcaption>Figure 1. Gel electrophoresis verification of the hpCA sequence after plasmid transformation into E. coli DH5𝛼. The primers used for colony PCR correspond to a 993bp-long product, which is close to the 1000bp bands observed.</figcaption>
   </figure>

Sequence and Features

Reference

Bury-Moné, S., Mendz, G. L., Ball, G. E., Thibonnier, M., Stingl, K., Ecobichon, C., Avé, P., Huerre, M., Labigne, A., Thiberge, J.-M., & DeReuse, H. (2008). Roles of α and β carbonic anhydrases of Helicobacter pylori in the urease-dependent response to acidity and in colonization of the murine gastric mucosa. Infection and Immunity, 76(2), 497-509. https://doi.org/10.1128/iai.00993-07

Chirica, L. C., Petersson, C., Hurtig, M., Jonsson, B.-H., Borén, T., & Lindskog, S. (2002). Expression and localization of α- and β-carbonic anhydrase in Helicobacter pylori. Biochimica Et Biophysica Acta (BBA) - Proteins and Proteomics, 1601(2), 192-199. https://doi.org/10.1016/s1570-9639(02)00467-3

Zhu, Y., Liu, Y., Ai, M., & Jia, X. (2022). Surface display of carbonic anhydrase on Escherichia coli for CO2 capture and mineralization. Synthetic and Systems Biotechnology, 7(1), 460-473. https://doi.org/10.1016/j.synbio.2021.11.008