Lucas H. Stevens

Subcellular localization of tryptophan decarboxylase, strictosidine synthase and strictosidine glucosidase in suspension cultured cells of Catharanthus roseus and Tabernaemontana divaricata.

The subcellular localization of tryptophan decarboxylase, strictosidine synthase and strictosidine glucosidase in suspension cultured cells of Catharanthus roseus (L.) G. Don and Tabernaemontana divaricata (L.) R. Br. ex Roem. et Schult, was investigated. It was found that tryptophan decarboxylase is an extra-vacuolar enzyme, whereas strictosidine synthase is active inside the vacuole. Strong indications were obtained for the localization of strictosidine glucosidase on the outside of the tonoplast. The results suggest that tryptamine is transported into the vacuole where it is condensed with secologanin to form strictosidine, and that strictosidine passes the tonoplast and is subsequently hydrolysed outside the vacuole.

Purification and characterization of strictosidine synthase from a suspension culture of Cinchona robusta

Abstract Four isoforms of strictosidine synthase (EC 4.3.3.2), which catalyses the stereospecific condensation of secologanin and tryptamine, were purified to h

Isolation and characterization of blackspot pigments from potato tubers

Blackspot related pigments were partially purified from bruised tubers of two commercial potato cultivars (cv. Bildtstar and cv. Lady Rosetta). Chemical characterization showed that these pigments consisted of protein and a relatively small amount of covalently bound constituents. These polymeric structures absorbed light throughout the visible spectrum without any maximum. They did not contain eumelanin. Quinic acid was detectable in hydrolysates of the pigments from Bildtstar, but not in those of Lady Rosetta, which indicated that chlorogenic acid may take part in blackspot formation, but is not essential for the discoloration. The results support the hypothesis that blackspot pigments are products of non-regulated reactions between nucleophilic amino acid residues in proteins, and quinones, which are derived from endogenous substrates of polyphenol oxidase. This means that blackspot formation most probably takes place in disintegrated cells.

Tyrosine and cysteine are substrates for blackspot synthesis in potato

Partially purified blackspot pigments from potato tubers (Solanum tuberosum L.) of two commercial cultivars were subjected to a microassay for melanin, which consisted of specific chemical degradation and subsequent HPLC analysis. Permanganate oxidation yielded pyrrole-2,3,5-tricarboxylic acid, whereas hydrolysis in hydriodic acid liberated aminohydroxyphenylalanine isomers. These results indicate that the polymeric pigments, which have previously been found to contain a protein matrix, carry crosslinked 5,6-dihydroxyindole-2-carboxylic acid and benzothiazine units. This leads to the conclusion that free tyrosine and free cysteine are incorporated in the proteinaceous pigments via the polyphenol oxidase catalysed pathway of melanogenesis in the process of blackspot formation. The findings are in accordance with the hypothesis that the process of blackspot formation is a non-regulated cascade of reactions in disintegrated tuber cells, rather than a finely tuned biosynthesis.