Pyrroloquinoline quinone dehydrogenases

Conversion of methanol into formaldehyde by methanol dehydrogenase. A complex array of genes is involved in this oxidation and the dehydrogenase contains pyrroloquinoline quinone (PQQ) as a cofactor (references in Ramamoorthi and Lidstrom 1995). Details of its function must, however, differ from that of methylamine dehydrogenase that also contains a quinoprotein—tryptophan tryptophylquinone (TTQ). 

Zayats M, Katz E, Baron R, Willner I. 2005. Reconstitution of apo-glucose dehydrogenase on pyrroloquinoline quinone-functionalized Au nanoparticles yields an electrically contacted hiocatalyst. J Am Chem Soc 127 12400-12406. 

Fructose dehydrogenase (FDH) having pyrroloquinoline quinone (PQQ) as a prosthetic group is an redox enzyme to catalyze the oxidation of fructose. A... 

Pyrroloquinoline quinone (PQQ) (or methoxatin) 6 is a coenzyme, responsible for the oxidation of methanol [7]. It has been found that cyclopropanol 4 inactivates the enzyme from M. methanica [8], the dimeric methanol dehydrogenase and the monomeric enzyme from a Pseudomonas PQQ-dependent methanol dehydrogenase [9] by forming adducts such as 7, through a one-electron oxidation process and the ready ring opening of a cyclopropyloxonium radical, Eq. (3) [8,9]. 

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. 

Shimao M, Tamogami T, Nishi K, Harayama S (1996) Cloning and characterization of the gene encoding pyrroloquinoline quinone-dependent poly(vinyl alcohol) dehydrogenase of Pseudomonas sp. strain VM15C. Biosci Biotechnol Biochem 60 1056-1062... 

Oxidation by direct H transfer from the a-carbon of alcohols to the pyrroloquinoline quinone (PQQ) cofactor of alcohol dehydrogenases was studied using ab initio quantum mechanical methods <2001JCC1732>. Energies and geometries were calculated at the 6-31G(d,p) level of theory, results were compared to available structural and spectroscopic data, and the role of calcium in the enzymatic reaction was explored. Transition state searches at the semi-empirical and STO-3G(d) level of theory provided evidence that direct transfer from the alcohol to C-5 of PQQ is energetically feasible. 

C. PQQ-Dependent Dehydrogenases (PQQ, Pyrroloquinoline Quinone) Properties of Mediators... 

Bacteria that oxidize methane or methanol (methylotrophs) employ a periplasmic methanol dehydrogenase that contains as a bound coenzyme, the pyrroloquinoline quinone designated PQQ or meth-oxatin (Eq. 15-51).442 I 1 1 This fluorescent ortfzo-quinone... 

ZnS colloids were also used by Kuwabata et al. to photoreduce C02 to formate [130]. In this system, the ZnS colloids also reduced pyrroloquinoline quinone which served as an electron mediator to the enzyme, methanol dehydrogenase, which could then reduce formate to methanol. In C02-saturated aqueous solution at pH 7 and under far-UV (280nm) illumination, quantum efficiencies of 7% and 6% were achieved for formate and methanol, respectively. 

M. Niculescu, R. Mieliauskiene, V. Laurinavicius and E. Csoregi, Simultaneous detection of ethanol, glucose and glycerol in wines using pyrroloquinoline quinone-dependent dehydrogenases based biosensors, Food Chem., 82(3) (2003) 481 89. 

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. 

Yoshida, H., Kojima, K., Witarto, A. B., and Sode, K. (1999). Engineering a chimeric pyrroloquinoline quinone glucose dehydrogenase improvement of EDTA tolerance, thermal stability and substrate specificity. Protein Eng., 12, 63-70. 

Mutzel, A., and Gorisch, H., 1991, Quinoprotein ethanol dehydrogenase preparation of the apo-form and reconstitution with pyrroloquinoline quinone and Ca " or Si ions. Agric. Biol. Chem. 55 172191726. 

Zheng, Y-J., and Bruice, T. C., 1997, Conformation of coenzyme pyrroloquinoline quinone and role of Ca in the catalytic mechanism of quinoprotein methanol dehydrogenase. Proc. Natl. Acad. Set 94 11881911886. 

The third case for likely enzymatic generation of an a-cyclopropyl radical or a cyclopropanone equivalent leading to target enzyme destruction is in the oxidation of cyclopropanol by certain bacterial alcohol dehydrogenases which have a novel type of redox coenzyme stoichiometrically bound and required for catalysis. The coenzyme has been termed pyrroloquinoline quinone (PQQ) (174) or methoxatin and the trivial name... 

Modified electrodes for this analytical purpose have mostly been formed by electrode adsorption of the mediator systems on the electrode surface or by electropolymerization [24,116]. Recently, for example, NAD(P)H oxidations have been performed on platinum or gold electrodes modified with a monolayer of pyrroloquinoline quinone (PQQ) [117] or on poly(methylene blue)-modified electrodes with different dehydrogenases entrapped in a Nafion film for the amperometric detection of glucose, lactate, malate, or ethanol [118]. In another approach, carbon paste electrodes doped with methylene green or meldola blue together with diaphorase were used for the NADH oxidation [119]. A poly(3-methylthio-phene) conducting polymer electrode was efficient for the oxidation of NADH [120]. By electropolymerization of poly(aniline) in the presence of poly(vinylsulfonate) counterions. 

An analogous amplification process for determination of phenols was proposed based on the kinetics of disappearance of /1-NADH reacting with quinone, which is derived from a phenol in a tyrosinase-catalyzed oxidation. LOD was as low as 50 nM in a 10 min assay . Amplification cycles were also achieved by combining a Pt electrode where phenols are oxidized with a polyurethane layer embedding pyrroloquinoline quinone-dependent glucose dehydrogenase, to catalyze the reduction of the oxidation products . 

Redox proteins that include quinone cofactor units play important roles in biological ET processes. Some of the quinoproteins include the quinone cofactor in a non-covalently linked configuration, such as the pyrroloquinoline quinone, PQQ, dependent enzymes, whereas other quinoproteins include the quinone cofactor covalently-linked to the protein, for example topaquinone (2,4,6-trihydroxyphenylalanine quinone, TPQ) dependent enzymes. A number of quinoproteins include in addition to the quinone cofactor an ET cofactor unit in another protein subunit. These cofactors may be metal ions or a cytochrome-type heme cofactor such as D-fructose dehydrogenase that is a heme containing PQQ-dependent enzyme. ... 

This method was applied to assemble integrated electrically-contacted NAD(P)-dcpcndcnt enzyme electrodes. The direct electrochemical reduction of NAD(l ) cofactors or the electrochemical oxidation of NAD(P)H cofactors are kineticaUy unfavored. Different diffusional redox mediators such as quinones, phenazine, phenoxazine, ferrocene or Os-complexes were employed as electrocatalysts for the oxidation of NAD(P)H cofactors An effective electrocatalyst for the oxidation of the NAD(P)H is pyrroloquinoline quinone, PQQ, (7), and its immobilization on electrode surfaces led to efficient electrocatalytic interfaces (particularly in the presence of Ca ions) for the oxidation of the NAD(P)H cofactors. This observation led to the organization of integrated electrically contacted enzyme-electrodes as depicted in Fig. 3-20 for the organization of a lactate dehydrogenase electrode. 

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