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It was demonstrated that an enzyme system(s) extracted from etiolated seedlings of Stizolobium hassjoo catalyzed the conversion of L-dihydroxyphenylalanine into stizolobinic acid. α-amino-6-carboxy-2-oxo-2H-pyran-3-propionic acid, and stizolobic acid, α-amino-6-earboxy-2-oxo-2H-pyran-4-propionic acid, in the presence of NADP+ or NAD+ under aerobic condition. Enzymically synthesized radioactive stizolobinic acid and stizolobic acid isolated from the reaction mixtures were purified and confirmed to have constant specific radioactivities by cocrystallization with authentic samples. Maximal activity of the enzyme preparation was obtained by using an insoluble polyphenol adsorbent (Polyclar AT) and a reducing agent (araboascorbic acid) in the extraction medium and by subsequent fractionation of the extract with ammonium sulfate followed by Sephadex G-25 gel filtration. Catalytic activity of the enzyme preparation was more unstable under aerobic condition than anaerobic, Attempts to stabilise the enzyme activity were made by the use of many substances which are known to stabilise other enzymes or expected to arrest the inactivation. Evidence is provided in this paper that the previously proposed biosynthetic pathways of stizolobinic acid and stizolobic acid from dihydroxyphenylalanine proceeded in the cell-free system from etiolated seedlings of S. hassjoo. [ABSTRACT FROM AUTHOR]

DL-3-Arsonoalanine has been synthesized by the Strecker synthesis from the unstable compound arsonoacetaldehyde. It inactivates pig heart cytosolic aspartate aminotransferase and inhibits aspartate ammonia-lyase by competing with aspartate (K/Km 0.23). The fumarate analogue (E)-3-arsonoacrylic acid and the malate analogue (RS)-3-arsonolactate also inhibit furnarate hydratase, competing with fumarate (K/Km 1.8) and malate (K/Km 1.6) respectively. Attempted non-enzymic transamination of 3-arsonoalanine gave elimination of arsenite, in contrast with the transamination of 3-phosphonoalanine, which is either successful or leads to loss of phosphate. [ABSTRACT FROM AUTHOR]

L-Phenylalanine ammonia-lyase (EC 4.3.1.5), which catalyses the first reaction in the biosynthesis of plant phenylpropanoid products, undergoes a transient increase in activity in excised sections of pea (Pisum sativum) epicotyl tissue. Exogenous supplies of pathway intermediates inhibit the initial development of phenylalanine ammonia-lyase activity and, if added at the time of high enzyme levels, cause a rapid decay in enzyme activity. The inhibitory effect of exogenous cinnamic acid, the itermediate product of the enzyme, is expressed very rapidly (lag < 1 h) with the initial rate of decay of enzyme activity and the final steady-state level of enzyme activity being dependent on the concentration of cinnamic acid. The response has been studied by density labelling in vivo with ²H from ²H2O with analysis of the equilibrium distribution of enzyme activity in high-resolution KBr density gradients. Cinnamic acid, at a concentration of 1 mM completely inhibits further incorporation of label and also markedly stimulates the removal of pre-existing, unlabelled enzyme. Inhibition of de novo production and stimulation of removal occur at concentrations of cinnamic acid as low as 10 μM. The initial increase in enzyme activity reflects an increase in the rate of de novo enzyme production against a low background rate of enzyme removal. The subsequent decay in enzyme activity is a result of a decline in production of the enzyme concomitant with a marked increase in the rate of removal of enzyme activity. In the presence of α aminooxy-β-phenylpropionic acid, which specifically inhibits phenylalanine ammonia-lyase and hence endogenous generation of cinnamic acid, high rates of enzyme production and low rates of enzyme removal are maintained leading to high levels of enzyme activity in the later stages of enzyme induction. These data indicate the operation of dual feedback modulation of phenylalanine ammonia-lyase in vivo following endogenous production of cinnamic acid. Such dual control by a product over both production and removal of an enzyme may be of general importance in slowly growing plant cells for the rapid removal of biosynthetic enzymes no longer required by the cell. [ABSTRACT FROM AUTHOR]

The level of N-acetyl-L-glutamate showed a diurnal rhythm, which was associated primarily with dietary ingestion. Since the hepatic level of arginine, activator for acetylglutamate, is one of the plausible principal factors responsible for the regulation of acetylglutamate level, the effect of arginine was further studied. The intraperitoneal administration of a large amount of arginine increased the level of acetylglutamate and the effect was augmented by the combined administration of glutamine. The arginine administration stimulated the hepatic synthesis of acetylglutamate as measured with [14C]glutamate as tracer, but did not affect the half-life (about 20 min) of the synthesized 14C-labeled acetylglutamate. The effect of arginine in raising the acetylglutamate level was also demonstrated hi vitro in isolated rat liver cells. On the other hand, pent-4-enoic acid, known to inhibit fatty acid oxidation and to decrease acetyl-CoA production in the liver, lowered the level of acetylglutamate in the rat liver cells. It was shown that the cellular ureogenic capacity, as measured with NH4Cl as nitrogen source, showed a close correlation with the varying levels of acetylglutamate, brought about by arginine or pent-4-enoic acid. The carbamoyl phosphate level, which may reflect the balance between its synthesis and utilization for citrulline synthesis, was low (about 1 nmol/g) and remained relatively constant in the livers of intact mice even when the rate of urea synthesis was expected to vary with changes in the dietary protein content. A transient increase in the level was brought about by agents expected to increase the synthesis or to inhibit the utilization, such as ammonia, N-acetylglutamate diethyl ester, or norvaline. These results, together with others, suggest that in intact animals the production of carbamoyl phosphate is coupled to its utilization for citrulline synthesis. [ABSTRACT FROM AUTHOR]