Oxidases o-diphenol

Dopa is converted by at least some insects into N-P-alanyldopamine, which is a preferred substrate for the o-diphenol oxidase of the insect pupal cuticle. Oxidation of this substrate plays a crucial role in the... 

Horigome and Kandatsu (63) prepared a Fuki (Petasites japon-icus miq. a traditional food plant in Japan) acetone-powder with a high o-diphenol oxidase activity. When the powder was added to a mixture of caffeic acid and casein, casein was gradually pigmented. A feeding test with rats demonstrated that there was a close relationship between the decrease in biological value of the pigmented casein and its color intensity. 

Monophenols are more slowly acting substrates as they have to be hydroxylated prior to then-oxidation to the corresponding o-quinones [7]. The commonest natural substrates for monophenol oxidase are probably tyrosine and p-coumaric acid or their derivatives. AU o-diphenol oxidases require the basic o-dihydroxyphenol structure for oxidase activity so that catechol is the simplest possible, but not necessarily the best, substrate 4-methyl catechol is usually the fastest [45]. The rate of oxidation of o-diphenols by PPO increases with increasing electron withdrawing power of substituents in the para position. o-Diphenol substitution (-CH3) at one of the positions adjacent to the -OH groups prevents oxidation. These positions should remain free for oxidation to take place [14]. 

Horigome, T. and Kandatsu, M. (1968). Biological value of protein allowed to react with phenolic compounds in presence of o-diphenol oxidase. Agric. Biol. Chem. 32 1093... 

Tyrosinase is an enzyme complex (phenolase, polyphenol oxidase are other names which have been used for this enzyme), which catalyses of the ortho hydroxylation of monohydric phenols. The enzyme, which should not be confused with L-tyrosine hydroxylase mentioned above, contains Cu (I) and catalyses two distinct reactions—the hydroxylation of monohydric phenols to o-diphenols (cresolase activity) and the oxidation of o-diphenols to o-quinones (catecholase or catechol oxidase activity) . Most enzymes of this type, which are widely distributed in both the plant and animal kingdoms, exhibit both cataljrtic functions. Thus typically, the conversion of L-tyrosine (5) to L-dopa (15) and dopaquinone (36) which occurs in melanin biosynthesis is catalysed by an enzyme of the tyrosinase category. The two activities appear, in the majority of cases, to be functions of the same enzyme. However, certain o-diphenol oxidases such as those from tea , sweet potato and tobacco have been reported to show no capacity to catalyse the hydroxylation reaction but this is most probably due to destruction of the cresolase activity during purification. 

FIGURE 8.11 A study of o-diphenol oxidase with catechol substrate including competitive (PHBA) and noncompetitive (phenylthiourea) inhibitors. (From Kimball, J., Enzyme kinetics, http / users.rcn.com/ jkimball.ma.ultranet/BiologyPages/E/EnzymeKinetics.html. With permission.)... 

Given the table of data below for the reaction of o-diphenol oxidase with catechol, draw the Lineweaver-Burk plot and find the slope of the graph and use the value of the y-intercept (1/Vinax) to obtain the Am value. Compare that value to the reciprocal of the x-intercept where (1/V) = 0. Submit your graph along with your calculations. We will need three significant figures to compare with the data in the next problem to see flie effect of an inhibitor. 

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