Pyrroloquinoline quinone enzymes with

Examples of surface-immobilized mediators are electropolymerized azines for electro-oxidation of The extreme form of this approach is formation of biocatalytic monolayer, comprising a surface-bound mediator species that is itself bound to a single enzyme molecule. Katz et al. report a complete cell based on novel architecture at both electrodes (Figure 7). On the anode side, the FAD center of glucose oxidase is removed from the enzyme shell and covalently attached to a pyrroloquinoline quinone (PQQ) mediator species previously immobilized on a gold surface. The GOx apoenzyme (enzyme with active center removed) is reintroduced in solution and selectively binds to FAD, resulting in a PQQ-... 

An additional condition may be imposed, even when a cofactor-independent enzyme is used, if a mediator molecule is involved in the electron transfer process, as is often the case with oxidases. Laccases, for example, may employ small-molecule diffusible mediator compounds in their redox cycle to shuttle electrons between the redox center of the enzyme and the substrate or electrode (Scheme 3.1) [1, 2]. Similarly, certain dehydrogenases utiHze pyrroloquinoline quinone. In biocatalytic systems, mediators based on metal complexes are often used. 

An alternative biosensor system has been developed by Hart et al. [44] which involves the use of the NAD+-dependent GDH enzyme. The first step of the reaction scheme involves the enzymatic reduction of NAD+ to NADH, which is bought about by the action of GDH on glucose. The analytical signal arises from the electrocatalytic oxidation of NADH back to NAD+ in the presence of the electrocatalyst Meldola s Blue (MB), at a potential of only 0Y. Biosensors utilising this mediator have been reviewed elsewhere [1,17]. Razumiene et al. [45] employed a similar system using both GDH and alcohol dehydrogenase with the cofactor pyrroloquinoline quinone (PQQ), the oxidation of which was mediated by a ferrocene derivative. 

Isolated oxidoreductases always depend on cofactors for the transfer of electrons. Enzyme groups which are well characterized with respect to their biochemistry are those requiring the nicotinamide coenzymes NAD or NADP, the flavins FAD or FMN and the ortho-quinoids such as pyrroloquinoline quinone (PQQ) or trihydroxy-phenylalanine (TOPA). 

Integrated electrically-contacted enzyme electrodes were prepared by the surface reconstitution of different apo-enzymes on electrode surfaces. The pyrroloquinoline quinone, PQQ, (7), was covalently linked to a cystamine monolayer associated with an Au-electrode, and A -(2-aminoethyl-FAD), (8), was covalently attached to the PQQ units. Fig. 3-4A. The integrated enzyme-electrode was then prepared by the reconstitution of apo-GOx on the FAD units. The surface coverage of the PQQ-FAD units was estimated to be 5.5x10 ° mole cnr. whereas the surface coverage of the reconstituted... 

Qjiinoproteins are a class of enzymes with cofactors possessing a quinone moiety, which participates directly in catalysis. Several dehydrogenases have been characterized which possess the dissociable cofactor 4,5-dihyro-4,5-dioxo-l-H-pyrrolo[2,3-f quinoline-2,7,9-tricarboxylic acid, which is now commonly called pyrroloquinoline quinone or PQCi. ° Earlier, several other enzymes were believed to possess covalently attached PQCJ as a cofactor. This is now known to be false and an unfortunate consequence of erroneous analytical techniques. Although there are no examples of enzymes possessing covalently bound PQft, covalent quinoproteins with other protein-derived quinones have been identified. 

A number of enzymes which catalyze oxidation reactions, including mammalian lysyl and plasma amine oxidases and bacterial alcohol dehydrogenases, have been determined to utilize pyrroloquinoline quinone (PQQ, methoxatin) as a cofactor (Duine et al., 1987). Substrates of the amine oxidases appear to be activated for a-proton abstraction by formation of a Schiff base with PQQ, fol-... 

An alternative way of NAD" recycling makes use of a three-enzyme cascade with molecular oxygen as the ultimate oxidant (Scheme 3.30) [339]. As in the methods described above, all the enzymes and cofactors have to be precipitated together. Thus, NADH which is produced by HLADH-catalyzed oxidation of a secondary alcohol is re-oxidized by diaphorase at the expense of pyrroloquinoline quinone (PQQ) [340]. The reduced form of the latter (PQQH2) is spontaneously oxidized by molecular oxygen producing hydrogen peroxide, which, in mm, is destroyed by catalase. 

Zhao, C., Wittstock, G. Scanning electrochemical microscopy for detection of biosensor and biochip surfaces with immobilized pyrroloquinoline quinone (PQQ)-dependent glucose dehydrogenase as enzyme label. Biosens Bioelectron 2005, 20, 1277-1284. 

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