The efficiency of immobilised glutamate oxidase decreases with surface enzyme loading: an electrostatic effect, and reversal by a polycation significantly enhances biosensor sensitivity.
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.
McMahon
Colm P
CP
UCD School of Chemistry and Chemical Biology, University College Dublin, Belfield, Dublin 4, Ireland.
Rocchitta
Gaia
G
Serra
Pier A
PA
Kirwan
Sarah M
SM
Lowry
John P
JP
O'Neill
Robert D
RD
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
D018698:Glutamic Acid; D011094:Polyethyleneimine; D000594:Amino Acid Oxidoreductases; C023061:L-glutamate oxidase