P-chip and P-chip bienzyme electrodes based on recombinant forms of horseradish peroxidase immobilized on gold electrodes.
Adsorption and bioelectrocatalytic activity of native horseradish peroxidase (HRP) and its recombinant forms on polycrystalline gold electrodes were studied. Recombinant forms of HRP were produced by a genetic engineering approach using an E. coli expression system. According to direct mass measurements with a quartz crystal microbalance, all the forms of HRP formed monolayer coverage of the enzyme on the gold surface. However, only gold electrodes modified with the recombinant HRP forms (non-glycosylated) exhibited high and stable current response to H2O2 due to its bioelectrocatalytic reduction based on direct electron transfer (ET) between gold and the active site of the enzyme. Introduction of a six-His tag either at the C-terminus or at the N-terminus of the enzyme molecule additionally increased the strength of the enzyme binding with the gold surface and the efficiency of direct ET. Immobilization of recombinant forms of HRP containing histidine functional groups on the surface of the gold electrode was used both for the development of a P-chip, a biosensor for hydrogen peroxide determination based on direct ET, and for the development of a bienzyme biosensor electrode for the determination of L-lysine based on co-immobilized recombinant forms of HRP and L-lysine-alpha-oxidase.
Ferapontova
E E
EE
Faculty of Pharmacy, Department of Analytical Chemistry, University of Alcala, Madrid, Spain. elena.ferapontova@uah.es
Grigorenko
V G
VG
Egorov
A M
AM
D000327:Adsorption; D000594:Amino Acid Oxidoreductases / Q000032:analysis* / Q000378:metabolism; D015374:Biosensing Techniques / Q000295:instrumentation*; D004563:Electrochemistry; D004579:Electron Transport / Q000502:physiology; D004800:Enzymes, Immobilized / Q000032:analysis* / Q000378:metabolism; D006046:Gold; D006735:Horseradish Peroxidase / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000378:metabolism; D008239:Lysine / Q000032:analysis* / Q000378:metabolism; D011791:Quartz / Q000032:analysis; D011994:Recombinant Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism
D004800:Enzymes, Immobilized; D011994:Recombinant Proteins; D011791:Quartz; D006046:Gold; D006861:Hydrogen Peroxide; D006735:Horseradish Peroxidase; D000594:Amino Acid Oxidoreductases; C041123:L-lysine oxidase; D008239:Lysine