title: The efficiency of immobilised glutamate oxidase decreases with surface enzyme loading: an electrostatic effect, and reversal by a polycation significantly enhances biosensor sensitivity.
abstract: The apparent Michaelis constant, K(M), for glutamate oxidase (GluOx) immobilised on Pt electrodes increased systematically with enzyme loading. The effect was due, at least in part, to electrostatic repulsion between neighbouring oxidase molecules and the anionic substrate, glutamate (Glu). This understanding has allowed us to increase the Glu sensitivity of GluOx-based amperometric biosensors in the linear response region (100+/-11 nA cm(-2)microM(-1) at pH 7.4; SD, n=23) by incorporating a polycation (polyethyleneimine, PEI) to counterbalance the polyanionic protein. Differences in the behaviour of glucose biosensors of a similar configuration highlight a limitation of using glucose oxidase as a model enzyme in biosensor design.
authors.affiliation: UCD School of Chemistry and Chemical Biology, University College Dublin, Belfield, Dublin 4, Ireland.
authors.forename: Colm P
authors.initials: CP
authors.lastname: McMahon
authors.forename: Gaia
authors.initials: G
authors.lastname: Rocchitta
authors.forename: Pier A
authors.initials: PA
authors.lastname: Serra
authors.forename: Sarah M
authors.initials: SM
authors.lastname: Kirwan
authors.forename: John P
authors.initials: JP
authors.lastname: Lowry
authors.forename: Robert D
authors.initials: RD
authors.lastname: O'Neill
mesh_terms: D000594:Amino Acid Oxidoreductases / Q000737:chemistry; D015374:Biosensing Techniques / Q000379:methods*; D018698:Glutamic Acid / Q000032:analysis*; D011094:Polyethyleneimine / Q000737:chemistry; D055672:Static Electricity; D013499:Surface Properties
chemical_list: D018698:Glutamic Acid; D011094:Polyethyleneimine; D000594:Amino Acid Oxidoreductases; C023061:L-glutamate oxidase