title: Enhanced activity of acetyl CoA synthetase adsorbed on smart microgel: an implication for precursor biosynthesis.
abstract: Acetyl coenzyme A (acetyl CoA) is an essential precursor molecule for synthesis of metabolites such as the polyketide-based drugs (tetracycline, mitharamycin, Zocor, etc.) fats, lipids, and cholesterol. Acetyl CoA synthetase (Acs) is one of the enzymes that catalyzes acetyl CoA synthesis, and this enzyme is essentially employed for continuous supply of the acetyl CoA for the production of these metabolites. To achieve reusable and a more robust entity of the enzyme, we carried out the immobilization of Acs on poly(N-isopropylacrylamide)-poly(ethylenimine) (PNIPAm-PEI) microgels via adsorption. Cationic PNIPAm-PEI microgel was synthesized by one-step graft copolymerization of NIPAm and N,N-methylene bis-acrylamide (MBA) from PEI. Adsorption studies of Acs on microgel indicated high binding of enzymes, with a maximum binding capacity of 286 μg/mg of microgel for Acs was achieved. The immobilized enzymes showed improved biocatalytic efficiency over free enzymes, beside this, the reaction parameters and circular dichroism (CD) spectroscopy studies indicated no significant changes in the enzyme structure after immobilization. This thoroughly characterized enzyme bioconjugate was further immobilized on an ultrathin membrane to assess the same reaction in flow through condition. Bioconjugate was covalently immobilized on a thin layer of preformed microgel support upon polyethylene terephthalate (PET) track etched membrane. The prepared membrane was used in a dead end filtration device to monitor the bioconversion efficiency and operational stability of cross-linked bioconjugate. The membrane reactor showed consistent operational stability and maintained >70% of initial activity after 7 consecutive operation cycles.
authors.affiliation: Department of Nanostructured Materials, Leibniz-Institut für Polymerforschung Dresden e. V. , Hohe Str. 6, D-01069 Dresden, Germany.
authors.forename: Nidhi Chandrama
authors.initials: NC
authors.lastname: Dubey
authors.forename: Bijay Prakash
authors.initials: BP
authors.lastname: Tripathi
authors.forename: Martin
authors.initials: M
authors.lastname: Müller
authors.forename: Manfred
authors.initials: M
authors.lastname: Stamm
authors.forename: Leonid
authors.initials: L
authors.lastname: Ionov
mesh_terms: D000106:Acetate-CoA Ligase / Q000737:chemistry* / Q000378:metabolism*; D000180:Acrylic Resins / Q000737:chemistry; D000327:Adsorption; D004800:Enzymes, Immobilized / Q000737:chemistry / Q000378:metabolism; D005782:Gels / Q000737:chemistry; D011094:Polyethyleneimine / Q000737:chemistry; D058105:Polymerization
chemical_list: D000180:Acrylic Resins; D004800:Enzymes, Immobilized; D005782:Gels; C052970:poly-N-isopropylacrylamide; D011094:Polyethyleneimine; D000106:Acetate-CoA Ligase
keywords: acetyl CoA synthesis; acetyl CoA synthetase; core−shell microgel; enzyme adsorption; smart polymer