CHEM 153C Lecture 9: Elucidation of Metabolic Pathways
8 Feb 2017
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Anabolic (biosynthetic): end product essential for growth: defect: organism cannot grow; accumulation of intermediates; feedback inhibition/repression. Catabolic (degradative): substrate serves as source of c, n for growth: defect: organism cannot grow; grow on alternative c, n source. Goals of elucidation: identify intermediates; identify/characterize enzymes (mechanism, location); characterize regulation. Problem in determining metabolic pathway: metabolites exist at low concentration at steady state relationship between molecules = mystery; need to change pathway/see impact. Powerful method: get mutant organism with defects in enzymes in pathway. Characteristics of metabolism: metabolites exist at low levels/steady state. To identify them, have to perturb steady-state or label them arising from precursors: metabolic inhibitors: Iodoacetic acid (ich2cooh): alkylating agent inhibits glycolytic glyceraldehyde-3-p-dehydrogenase by alkylating enzyme (precursor of. Shows precursor-product relationships: labeled a would label b, c, d, e (whatever comes after. Labeling is sensitive tells origin of intermediates: common radioisotopes: 3h, 14c, 32p, 35s. Stable isotopes: 13c, 15n: detection of both isotopes:
