title: Probing mechanisms for enzymatic activity enhancement of organophosphorus hydrolase in functionalized mesoporous silica.
abstract: We have previously reported that organophosphorus hydrolase (OPH) can be spontaneously entrapped in functionalized mesoporous silica (FMS) with HOOC- as the functional groups and the entrapped OPH in HOOC-FMS showed enhanced enzyme specific activity. This work is to study the mechanisms that why OPH entrapped in FMS displayed the enhanced activity in views of OPH-FMS interactions using spectroscopic methods. The circular dichroism (CD) spectra show that, comparing to the secondary structure of OPH free in solution, OPH in HOOC-FMS displayed increased alpha-helix/beta-strand transition of OPH with increased OPH loading density. The fluorescence emission spectra of Trp residues were used to assess the tertiary structural changes of the enzyme. There was a 42% increase in fluorescence. This is in agreement with the fact that the fluorescence intensity of OPH was increased accompanying with the increased OPH activity when decreasing urea concentrations in solution. The steady-state anisotropy was increased after OPH entrapping in HOOC-FMS comparing to the free OPH in solution, indicating that protein mobility was reduced upon entrapment. The solvent accessibility of Trp residues of OPH was probed by using acrylamide as a collisional quencher. Trp residues of OPH-FMS had less solvent exposure comparing with free OPH in solution due to its electrostatical binding to HOOC-FMS thereby displaying the increased fluorescence intensity. These results suggest the interactions of OPH with HOOC-FMS resulted in the protein immobilization and a favorable conformational change for OPH in the crowded confinement space and accordingly the enhanced activity.
authors.affiliation: Pacific Northwest National Laboratory, Richland, WA 99352, USA.
authors.forename: Baowei
authors.initials: B
authors.lastname: Chen
authors.forename: Chenghong
authors.initials: C
authors.lastname: Lei
authors.forename: Yongsoon
authors.initials: Y
authors.lastname: Shin
authors.forename: Jun
authors.initials: J
authors.lastname: Liu
mesh_terms: D043303:Aryldialkylphosphatase / Q000737:chemistry*; D002384:Catalysis; D002942:Circular Dichroism; D004800:Enzymes, Immobilized / Q000737:chemistry*; D016062:Porosity; D017433:Protein Structure, Secondary; D012822:Silicon Dioxide / Q000737:chemistry*; D013050:Spectrometry, Fluorescence
chemical_list: D004800:Enzymes, Immobilized; D012822:Silicon Dioxide; D043303:Aryldialkylphosphatase