title: Cross-linked polymer nanofibers for hyperthermophilic enzyme immobilization: approaches to improve enzyme performance.
abstract: We report an enzyme immobilization method effective at elevated temperatures (up to 105 °C) and sufficiently robust for hyperthermophilic enzymes. Using a model hyperthermophilic enzyme, α-galactosidase from Thermotoga maritima, immobilization within chemically cross-linked poly(vinyl alcohol) (PVA) nanofibers to provide high specific surface area is achieved by (1) electrospinning a blend of a PVA and enzyme and (2) chemically cross-linking the polymer to entrap the enzyme within a water insoluble PVA fiber. The resulting enzyme-loaded nanofibers are water-insoluble at elevated temperatures, and enzyme leaching is not observed, indicating that the cross-linking effectively immobilizes the enzyme within the fibers. Upon immobilization, the enzyme retains its hyperthermophilic nature and shows improved thermal stability indicated by a 5.5-fold increase in apparent half-life at 90 °C, but with a significant decrease in apparent activity. The loss in apparent activity is attributed to enzyme deactivation and mass transfer limitations. Improvements in the apparent activity can be achieved by incorporating a cryoprotectant during immobilization to prevent enzyme deactivation. For example, immobilization in the presence of trehalose improved the apparent activity by 10-fold. Minimizing the mat thickness to reduce interfiber diffusion was a simple and effective method to further improve the performance of the immobilized enzyme.
authors: Department of Chemical and Biomolecular Engineering North Carolina State University , Raleigh, North Carolina 27695, United States.
authors: Christina
authors: C
authors: Tang
authors: Carl D
authors: CD
authors: Saquing
authors: Stephen W
authors: SW
authors: Morton
authors: Brittany N
authors: BN
authors: Glatz
authors: Robert M
authors: RM
authors: Kelly
authors: Saad A
authors: SA
authors: Khan
mesh_terms: D002384:Catalysis; D003432:Cross-Linking Reagents / Q000737:chemistry*; D004563:Electrochemistry; D004800:Enzymes, Immobilized / Q000737:chemistry*; D004926:Escherichia coli / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008855:Microscopy, Electron, Scanning; D057139:Nanofibers / Q000737:chemistry*; D036103:Nanotechnology / Q000379:methods*; D011108:Polymers / Q000737:chemistry*; D011142:Polyvinyl Alcohol / Q000737:chemistry; D012997:Solvents / Q000737:chemistry; D013696:Temperature; D020124:Thermotoga maritima / Q000378:metabolism
chemical_list: D003432:Cross-Linking Reagents; D004800:Enzymes, Immobilized; D011108:Polymers; D012997:Solvents; D011142:Polyvinyl Alcohol