We propose to continue our biochemical studies of the metabolism of several aromatic cyanogenic glycosides, natural products, which occur in cyanogenic plants. The studies are desgined to obtain fundamental biochemical information concerning the metabolism of natural products on the assumption that any principles established will be applicable to other such compounds. Plant natural products include many substances (alkaloids, coumarins, flavonoids, glucosinolates) which are of medicinal and toxicological interest. Aromatic cyanogenic glycosides are derived from L-tyrosine and L-phenylalanine by a biosynthetic sequence involving the corresponding N-hydroxyamino acid, aldoxime, nitrile and cyanoydrin. Two of the four steps are catalyzed by mixed-function oxygenases. The biosynthetic sequence catalyzed by sorghum microsomes is metabolically channeled so that intermediates produced in situ are used more effectively than the same intermediates added externally to the microsomes. We propose to see whether particules from other cyanogenic seedlings (vetch and flax) are similarly channeled. Because efforts to solubilize the several enzymes involved have not yielded a soluble, purified protein whose molecular weight (size) could be determined, we shall used the technique of radiation inactivation to measure the molecular weight of the enzyme(s) involved. Such information should indicate whether the several enzymes are separate catalytic entitles or whether they function as a multi-enzyme complex. Cyanogenic sorghum seedlings possess an effective biosynthetic system for producing tyrosine from starch. Since little is known about the enzymes involved or their regulation, we propose to study the shikimate pathway in such seedlings. In particular, we will examine the role of the chloroplast in this process, since this organelle is now recognized as a site of synthesis of amino acids in plants. Finally, the mechanism of putative dehydration reaction (aldoxime greater than nitrile + H2O) which appears to require NADPH will be studied.
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