title: Enhancing bioplastic-substrate interaction via pore induction and directed migration of enzyme location.
abstract: We demonstrate two novel approaches to enhance interactions of polymer-immobilized biomolecules with their substrates. In the first approach, diisopropylfluorophosphatase (DFPase) containing poly(urethane) (PU) coatings were made microporous by incorporating, then extracting, poly(ethylene glycol)-based diesters as porogens. Incorporation of 2% w/w porogen increased the effective diffusion coefficient of diisopropylfluorophosphate (DFP) through the coatings by 30% and increased the apparent turnover number of immobilized DFPase 3-fold. In the second approach, prior to immobilization, hydrophobic modification of DFPase was achieved through its conjugation with a dimer/trimer mixture of a uretdione based on 1,6-diisocyanatohexane. When the hydrophobically modified DFPase was immobilized in coatings, catalytic activity was 4-fold higher than that of the equivalent, immobilized, native DFPase. This activity enhancement was independent of the presence or absence of pores. Confocal microscopy images of coatings containing fluorescently labeled lysozyme show that the native enzyme is distributed uniformly over the entire thickness of the coatings. Hydrophobically modified and fluorescently labeled lysozyme is accumulated only in the upper 10 microm cross-sectional layer of a 100 microm-thick coating. Interactions of bioplastics with their substrates are tunable either by pore induction in a polymer or by directed migration of the hydrophobically modified biomolecule to the desired location. The latter approach has broad implications, including overcoming mass transfer limitations experienced by immobilized biocatalysts.
authors.affiliation: Department of Bioengineering and McGowan Institute for Regenerative Medicine, Center for Biotechnology and Bioengineering, University of Pittsburgh, Pennsylvania 15219, USA.
authors.forename: Bhalchandra S
authors.initials: BS
authors.lastname: Lele
authors.forename: Glenn
authors.initials: G
authors.lastname: Papworth
authors.forename: Vicky
authors.initials: V
authors.lastname: Katsemi
authors.forename: Heinz
authors.initials: H
authors.lastname: Rüterjans
authors.forename: Igor
authors.initials: I
authors.lastname: Martyano
authors.forename: Kenneth J
authors.initials: KJ
authors.lastname: Klabunde
authors.forename: Alan J
authors.initials: AJ
authors.lastname: Russell
mesh_terms: D020099:Coated Materials, Biocompatible / Q000138:chemical synthesis*; D004058:Diffusion; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000737:chemistry*; D005456:Fluorescent Dyes; D018613:Microscopy, Confocal; D008855:Microscopy, Electron, Scanning; D009113:Muramidase / Q000378:metabolism; D044345:Phosphoric Triester Hydrolases / Q000737:chemistry*; D011092:Polyethylene Glycols / Q000737:chemistry; D011140:Polyurethanes / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013379:Substrate Specificity
chemical_list: D020099:Coated Materials, Biocompatible; D004800:Enzymes, Immobilized; D005456:Fluorescent Dyes; D011140:Polyurethanes; D011994:Recombinant Proteins; D011092:Polyethylene Glycols; D044345:Phosphoric Triester Hydrolases; C020061:diisopropyl-fluorophosphatase; D009113:Muramidase