Flavoenzymes

F. Muller (Ed.), Chemistry and Biochemistry of Flavoenzymes, CRC Press, Boca Raton, 1991. 

FIGURE 6-9 Electrical contact of a flavoenzyme by its reconstitution with a relay-FAD semisynthetic cofactor. Fc = ferrocene. (Reproduced with permission from reference 2.)... 

Catalysis by flavoenzymes has been reviewed and various analogues of FAD have been prepared e.g. P -adenosine-P -riboflavin triphosphate and flavin-nicotinamide dinucleotide ) which show little enzymic activity. The kinetic constants of the interaction between nicotinamide-4-methyl-5-acetylimidazole dinucleotide (39) and lactic dehydrogenase suggest the presence of an anionic group near the adenine residue at the coenzyme binding site of the enzyme. ... 

Salicylate is an intermediate in the metabolism of PAHs including naphthalene and phen-anthrene, and its degradation involves oxidation to catechol. The hydroxylase (monooxygenase) has been extensively studied (references in White-Stevens and Kamin 1972) and in the presence of an analog that does not serve as a substrate, NADH is oxidized with the production of H2O2 (White-Stevens and Kamin 1972). This uncoupling is characteristic of flavoenzymes and is exemplified also by the chlorophenol hydroxylase from an Azotobacter sp. that is noted later. 

A group of flavoenzyme reductases mediate the reduction both nitrate esters and C= C double bonds generally activated with carbonyl or nitro functions. The structures of the substrates vary widely (references in Faber 1997). 

Latham J, AC Walsh (1986) Retention of configuration in oxidation of a chiral boronic acid by the flavoenzyme cyclohexanone oxygenase. J Chem Soc Chem Commun 527-528. 

Walsh and coworkers oxidized ethyl p-tolyl sulfide on an analytical scale to the S-sulfoxide of 64% enantiomeric purity using a bacterial flavoenzyme cyclohexanone monooxygenase derived from Adnetobacter . Using a flavin adenine dinucleotide containing monooxygenase purified from hog liver microsomes yielded the R-sulfoxide of 90% enantiomeric purity. HPLC on a column containing a 3,5-dinitrobenzoyl-D-phenylglycine chiral stationary phase was used to determine the optical purity of the sulfoxides. 

Stereoselective oxygen transfer to the sulphur atom of alkyl aryl sulphides catalyzed by 2-flavoenzyme monooxygenases afforded optically active sulphoxides in high optical yields . For instance, with ethyl p-tolyl sulphide as substrate cyclohexanone monooxygenase from Actinetobacter produces predominantly (— )-(S)-sulphoxide with 64% e.e. In contrast, FAD-containing dimethylaniline monooxygenase purified from hog liver microsomes affords (+ )-(i )-enantiomer of this sulphoxide with 90% optical purity . ... 

Glutathione reductase (GR) catalyzes the reduction of oxidized glutathione (GSSG) to reduced glutathione (GSH) using NADPH provided from the hexose monophosphate pathway. GR, a ubiquitous flavoenzyme, maintains a high value of two for the GSH/GSSG ratio in the red blood cells. l,3-Bis(2-chloroethyl)-nitrosourea (BCNU) selectively inhibits cellular GR. GR is composed of two identical subunits, each of molecular mass 50 kDa (S8). The three-dimensional structure and mechanism of catalysis have been established for human GR (K17). 

Another thermophilic flavin reductase, flavoenzyme Bfl, from Bacillus sp. DSM411 was reported to have high thermostability [167], compared to the DszD from R. erythropolis D-l or the P. Polymyxa A-l. It had an optimum temperature of 70°C and optimum pH of 6.0. It retained 90% of its activity at 70°C after 30 min of incubation and was stable in pH range 2-12. 

Chordate neurons (early Cambrian) As above plus first hydroxylations giving serotonin and dopamine iron/pterin chemistry in cytoplasm vesicle filled in centre of cell Recovery by amine oxidation (flavoenzymes)... 

Hemmerich, P. The present status of flavin and flavoenzyme chemistry. In Progress in the chemistry of anorganic natural products 33,451 (1976)... 

FIGURE 33 Electrical contacting of a flavoenzyme by its reconstitution with a relay-FAD semisynthetic cofactor. 

The VAPOR enzymes are flavoenzymes and can be isolated from thermophilic bacilli. They are especially valuable because they allow the regeneration of all four forms of the pyridine nucleotides NADH, NAD+ NAD PH, NADP+ according to the following equations [55,61] ... 

The conversion (19) of thiols to disulphides coupled with reduction of flavin (vitamin B2 family) is a topic of import in connection with coenzyme reactivity in flavoenzymes. Since flavin oxidation of thiols involves nucleophilic attack of thiolate ion in the rate-determining step (Loechler and Hollocher, 1975 Yokoe and Bruice, 1975), this biologically important reaction would be markedly affected by hydrophobic environments. 

Flavin oxidation of carbanions has also been of much concern since active intermediates in some flavoenzyme-mediated reactions (amino acid oxidase, lactate oxidase, etc.) are carbanions (Kosman, 1977). Flavin oxidation of nitroethane carbanion (20), which had not been achieved in non-enzymatic systems, occurs with [56] bound to CTAB micelles (Shinkai etal., 1976b). This suggests that the nitroethane carbanion is also activated by the micellar environment. 

Attempts to reduce interference and minimize the effect of variations in oxygen tension have resulted in the development of biosensors with improved linear ranges which operate at lower electrode potentials. They incorporate artificial electron acceptors, called mediators, to transfer electrons from the flavoenzyme (e.g. glucose oxidase) to the electrode and thus are not dependent on oxygen. Ferrocene (bis(i75-cyclopentadienyl)iron) and its derivatives are examples of redox mediators for flavoenzymes. The reaction now becomes... 

Light DR, Waxman DJ, Walsh C. 1982. Studies on the chirality of sulfoxidation catalyzed by bacterial flavoenzyme cyclohexanone monooxygenase and hog liver flavin adenine dinucleotide containing monooxygenase. Biochemistry 21 2490-2498. 

Qrunones can accept one or two electrons to form the semiquinone anion (Q ") and the hydroquinone dianion (Q ). Single-electron reduction of a quinone is catalyzed by flavoenzymes with relatively low substrate selectivity (Kappus, 1986), for instance NADPH cytochrome P-450 reductase (E.C. 1.6.2.3), NADPH cytochrome b5 reductase (E.C. 1.6.2.2), and NADPH ubiquinone oxidoreductase (E.C. 1.6.5.3). The rate of reduction depends on several interrelated chemical properties of a quinone, including the single-electron reduction potential, as well as the number, position, and chemical characteristics of the substituent(s). The flavoenzyme DT-diphorase (NAD(P)H quinone acceptor oxidoreductase E.C. 1.6.99.2) catalyzes the two-electron reduction of a quinone to a hydroquinone. 

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