In situ kinetic measurements of d-amino acid oxidase in rat liver with respect to its substrate specificity

Abstract

D-Amino acid oxidase activity was demonstrated in peroxisomes of rat liver using unfixed cryostat sections and a histochemical technique using cerium ions as capture reagent for hydrogen peroxide and diaminobenzidine, cobalt ions and exogenous hydrogen peroxide to visualize the final reaction product for light microscopical analysis. Cytophotometric analysis of liver sections revealed similar zero-order reaction velocities of D-amino acid oxidase with activity twice as high in periportal areas as in pericentral areas of liver lobuli when using either D-proline or D,L-thiazolidine-2-carboxylic acid as substrates. On the other hand, a 4-5 times higher KM value was found for D-proline than for D,L-thiazolidine-2-carboxylic acid. The KM values in periportal and pericentral areas were similar for each substrate. These findings support the suggestion that the physiological substrate for D-amino acid oxidase may be D,L-thiazolidine-2-carboxylic acid, the adduct of cysteamine and glyoxylic acid. D-Amino acid oxidase may play a role in vivo in the production of oxalate which may participate in metabolic control processes as an intracellular messenger molecule.

In situ heterogeneity of peroxisomal oxidase activities: An update

“…The protein was detected in peroxisomes of liver (Veenhuis & Wendelaar Bonga, 1979;Stefanini et al, 1985;Perotti et al, 1987, kidney (Veenhuis andWendelaar Bonga, 1977;Usuda et al, 1986Usuda et al, , 1991Perotti et al, 1987;Yokota et al, 1987;Angermiiller, 1989), brain (Arnold et al, 1977(Arnold et al, ,1979, and in yeast (Veenhuis et al, 1976) (Table 2). 5) seems to be the physiological substrate for this enzyme (Hamilton et al, 1979, Frederiks et al, 1993a and the adduct is considered to be an important effector of some enzyme reactions, the enzyme may also be involved in metabolic processes, such as the intracellular regulation of oxalate levels, the release of histamine, the effects of nicotine on the metabolism, control of cell growth, and effects on several intracellular messenger systems that transduce signals of hormones, especially insulin (Hamilton et al, 1979(Hamilton et al, , 1987Hamilton, 1985;Skorczynski & Hamilton, 1986). Although this enzyme is found in a wide variety of organisms (Blaschko & Hawkins, 1952;Kilby & Neville, 1957;Wohlrab, 1965;Veenhuis et aI., 1976;Veenhuis & Wendelaar Bonga, 1977;Konno et al, 1982), the physiological function of a D-amino acid oxidizing system in higher animals is still a matter of debate, since D-amino acids are only found in some classes of bacteria and some insects and worms, and not in higher animals (D'Aniello et al, 1993).…”

Extinction coefficient of polymerized diaminobenzidine complexed with cobalt as final reaction product of histochemical oxidase reactions

“…It indicates that the cerium-DAB-cobalt-hydrogen peroxide method is much more sensitive than the tetrazolium salt method in detecting oxidase activity at the light microscopic level. This may explain why the enzyme could not be demonstrated in rat liver with the tetrazolium salt method (Kooij et al, unpublished results), but was detected with the cerium salt procedure (Angermtiller and Fahimi 1988 a,b;Patel et al 1991;Frederiks et al 1993). Rat liver has a much lower D-amino acid oxidase activity than rat kidney (Angermtiller and Fahimi 1988 a,b).…”

“…The cerium salt procedure was performed as described previously Frederiks et al 1993). The incubation medium consisted of 300 mM TRIS-maleate buffer, pH 7.8, 30 mM cerium chloride (Merck, Darmstadt, Germany), 100 mM sodium azide and 20 mM D-proline.…”

“…However, we were not able to produce the polyDAB-cobalt complex in vitro. Therefore, D-amino acid oxidase activity was analyzed quantitatively in proximal tubular cells in serial sections of rat kidney as detected with the tetrazolium salt method and with the cerium-DAB-cobalt-hydrogen peroxide method (Gossrau et al 1989;Patel et al 1991;Frederiks et al 1993). The validity of both reactions for quantitative purposes and the known extinction coefficient of tetranitro BT-formazan (Van Noorden 1984) enabled calculation of this coefficient for the polyDAB-cobalt complex.…”

The Origin of the Differences in (R)‐ and (S)‐Desmethyldesferrithiocin: Iron‐Clearing Propertiesa

“…38–39 We therefore elected to examine whether DMDFT, a thiazoline carboxylic acid, or its ferric chelates might affect the activity of d‐amino acid oxidase (d‐AAO). Thiazolidine carboxylates have been reported to be excellent substrates of d‐AAO, 40–41 a peroxide‐producing enzyme present in high concentrations in the peroxisomes of mammalian liver and kidney proximal tubules. 36 Moreover, d‐AAO‐catalyzed metabolism of a d‐cysteine derivative has been implicated in acute nephrotoxicity in the dog and results in selective ultrastructural changes in the S1 and S2 cells of the proximal tubules and decreased renal clearance.…”