title: Periplasmic expression of carbonic anhydrase in Escherichia coli: a new biocatalyst for CO(2) hydration.
abstract: Carbonic anhydrase is a valuable and efficient catalyst for CO(2) hydration. Most often the free enzyme is employed which complicates catalyst recycling, and can increase cost due to the need for protein purification. Immobilization of the enzyme may address these shortcomings. Here we report the development of whole-cell biocatalysts for CO(2) hydration via periplasmic expression of two forms of carbonic anhydrase in Escherichia coli using two different targeting sequences. The enzymatic turnover numbers (kcat ) and catalytic efficiencies (k(cat)/K(M)) were decreased by an order of magnitude as compared to the free soluble enzyme, indicating the introduction of transport limitations. However, the thermal stabilities were improved for most configurations (>88% activity retention up to 95°C for three of four whole-cell biocatalysts), operational stabilities were more than satisfactory (100% retention after 24 h of use for all four whole-cell biocatalysts), and CO(2) hydration was significantly enhanced relative to the uncatalyzed reaction (~50-70% increase in CaCO(3) precipitate formed). A significant advantage of the whole-cell approach is that protein purification is no longer necessary, and the cells can be easily separated and recycled in future applications including biofuel production, biosensors, and carbon capture and storage.
authors: Department of Chemical Engineering, Columbia University, 500 W 120th Street, New York, NY 10027, USA.
authors: Tushar N
authors: TN
authors: Patel
authors: Ah-Hyung Alissa
authors: AH
authors: Park
authors: Scott
authors: S
authors: Banta
mesh_terms: D002245:Carbon Dioxide / Q000378:metabolism; D002256:Carbonic Anhydrases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D015870:Gene Expression; D007700:Kinetics; D019897:Periplasm / Q000201:enzymology*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013696:Temperature
chemical_list: D011994:Recombinant Proteins; D002245:Carbon Dioxide; D002256:Carbonic Anhydrases