_L-Proline sensor based on layer-by-layer immobilization of thermostable dye-linked L-proline dehydrogenase and polymerized mediator

Abstract

L-Proline sensor has been fabricated through multilayer assembly of recombinant dye-linked L-proline dehydrogenase (L-proDH) originally from hyperthermophilic archaeon (Therococcus profundus) and polymerized mediator, poly(allylamine) ferrocene (PAA-Fc), on a gold electrode by electrostatic layer-by-layer alternate adsorption method. The characteristic redox peaks of ferrocene groups were obvious exhibited in cyclic voltammograms, and both anodic and cathodic peaks increased with PAA-Fc/L-proDH bilayer number which confirmed the formation of a multilayer structure. Further experiments showed that an electrode fabricated in this way catalyzed the oxidation of L-proline. It suggested that polymerized mediator could transfer electrons between electrode surface and immobilized L-proDH. Ampeormetric experiments showed that the current response to L-proline increased with PAA-Fc/L-proDH bilayer number. The stability of the sensor was measured, and it kept a 40% relative response after 30 days storage in buffer under 4 8C. q

Gene Expression and Characterization of a Third Type of Dye-LinkedL-Proline Dehydrogenase from the Aerobic Hyperthermophilic Archaeon,Aeropyrum pernix

“…7) We have succeeded in developing an application fortype LPDH in an electrochemical sensor used to assay L-proline. 8) These enzymes are highly stable and totally novel flavin-containing dehydrogenases.…”

Construction of L‐Lysine Sensor by Layer‐by‐Layer Adsorption of L‐Lysine 6‐Dehydrogenase and Ferrocene‐Labeled High Molecular Weight Coenzyme Derivative on Gold Electrode

“…It has already been proved that the LBL technique was an efficient method to immobilize enzymes, which was crucial in the construction of amperometric biosensors [19 -22]. In our previous works, the LBL method has been utilized to immobilize enzymes on electrode surface [23,24], and in the present research, we expended this method to the construction of reagentless l-lysine biosensor.…”

“…The electrode was finely polished and electrochemically cleaned by cyclic voltammetry between À 0.2 to þ 1.7 V vs. Ag/AgCl in 1 M H 2 SO 4 at a scan rate of 100 mV s À1 until a reproducible plot was obtained. The surface roughness was calculated at 1.6 according to our previous work [24]. Bare Au electrode after pretreated was immersed in 10 mM 3-MPS aqueous solutions for 12 h, and thoroughly washed by water.…”

Porphyrin “Mille-Feuilles” photo-electrodes

“…3a). This is a voltammetric response of surface bound species for which there is instant charge compensation by ions from the organic phase [24][25][26][27][28]. It is interesting to notice that if the forward peak potential remains constant, the backward peak potential shifts upon increasing scan rates. )…”