Starting materials:Amino acids and nucleic acidspseudopelletierinenicotinecaffeineergotmorphinecocaineornithinepurinetyrosinelysinetryptophanAlkaloid Biosynthesis:Nitrogen containing compoundsS. OConnorBiochemistry 7042/5/08ajmalinetryptophan

Endogenous Roles of Natural ProductsinsecticideattractionsignallingUV-protectantsantibiotic/antifungalscentcolorS. OConnorBiochemistry 7042/5/08

Endogenous Roles of Natural Productsalkaloids made by plants

fungi

bacteriaS. OConnorBiochemistry 7042/5/08ergotcocaine

Tetrahydroisoquinoline BiosynthesisS. OConnorBiochemistry 7042/5/08

S. OConnorBiochemistry 7042/5/08How are new genes discovered??Genome Mining (for sequenced prokaryotes/fungi)

Reverse genetics

Homology cloning (P450s, acetyltransferases, terpene synthases)

Large scale sequencingseqeunce all (Taxol)sequence cell type (tabersonine OMe transferase- next page)

Functional expression

6.RNAi screens

S. OConnorBiochemistry 7042/5/08How are new genes discovered??expressed in leaf epidermis

Tetrahydroisoquinoline BiosynthesisS. OConnorBiochemistry 7042/5/08J Nat Prod (2005), 68, 666-673.

How determine the order of the steps in a pathway?

track biosynthetic intermediates before and after knocking out genes

assay enzymes in vitro with different substratesS. OConnorBiochemistry 7042/5/08J Nat Prod (2005), 68, 666-673.

Tetrahydroisoquinoline BiosynthesisTwo oxidative pathsS. OConnorBiochemistry 7042/5/08

Tetrahydroisoquinoline BiosynthesisS. OConnorBiochemistry 7042/5/08

Tetrahydroisoquinoline BiosynthesisS. OConnorBiochemistry 7042/5/08JBC (1995) 270 24475-24481

Tetrahydroisoquinoline BiosynthesisS. OConnorBiochemistry 7042/5/08

Tetrahydroisoquinoline Biosynthesis

STSP450 enzymeS. OConnorBiochemistry 7042/5/08Phytochem 1993 32 79

Tetrahydroisoquinoline BiosynthesisS. OConnorBiochemistry 7042/5/08Tetrahedron Lett 1994 35 3897

S. OConnorBiochemistry 7042/5/08Tetrahydroisoquinoline BiosynthesisTrends in Biotechnology (2005), 23(7), 331-333.

S. OConnorBiochemistry 7042/5/08Tetrahydroisoquinoline Biosynthesis3400 BCjoy plant by Sumerians

1895 Heinrich Dreser working for The Bayer Company

Immunolocalization used to track location of enzymes in morphine biosynthesisS. OConnorBiochemistry 7042/5/08Tetrahydroisoquinoline Biosynthesis

Curr Opin Plant Biol 2005 8 292S. OConnorBiochemistry 7042/5/08Tetrahydroisoquinoline Biosynthesis

Immunolocalization used to track location of enzymes in morphine biosynthesis

S. OConnorBiochemistry 7042/5/08PNAS 2004 101 14091PNAS 2005 102 8495Human Morphine Biosynthesis

we have an opiate receptor

thought to bind peptidesYPFP (Science 1981 212 75)

Caffeinetheobrominexanthosine7-methylxanthosine7-methylxanthineS. OConnorBiochemistry 7042/5/08Caffeine BiosynthesisPurine

Trends Plant Sci. (2001) 6 407.S. OConnorBiochemistry 7042/5/08Caffeine BiosynthesisPurine

Genetically engineered decaf coffee

70% reduction in caffeine content by using RNAi of theobromine synthaseotherwise normal phenotypeNature (2003) 423, 823

Caffeine appears to act as insecticideTrans. Res 2006

Recently discovered natural variant deficient in caffeine synthase Nature (2004) 429, 826.

Promoter of the 3 methyl transferases recently discoveredJ. Biotech. (2005) 119, 20-25.S. OConnorBiochemistry 7042/5/08Caffeine Biosynthesis

EngineeringCaffeinetheobrominexanthosine7-methylxanthosine7-methylxanthine

Ergot Alkaloidstryptophan

S. OConnorBiochemistry 7042/5/08Plant Med 2006

Ergot Alkaloidstryptophan

Mol. Gen. Gen. (1999) 261, 133-141The Alkaloids, G. Cordell pp. 170-218S. OConnorBiochemistry 7042/5/08

Ergot AlkaloidstryptophanS. OConnorBiochemistry 7042/5/08Mol. Gen. Gen. (1999) 261, 133-141The Alkaloids, G. Cordell pp. 170-218

Journal of Biological Chemistry (1995), 270(41), 24475-81. Mol. Gen. Gen. (1999) 261, 133-141The Alkaloids, G. Cordell pp. 170-218Potential for combinatorial biosynthesis?Central amino acid varies

Ergovaline eliminated from a strainof symbiantryegrass staggers(PNAS (2001) 98 12820)S. OConnorBiochemistry 7042/5/08Ergot Alkaloidstryptophan

Mol. Gen. Gen. (1999) 261, 133-141The Alkaloids, G. Cordell pp. 170-218S. OConnorBiochemistry 7042/5/08Ergot Alkaloidstryptophan

S. OConnorBiochemistry 7042/5/08Terpene Indole Alkaloidstryptophan

S. OConnorBiochemistry 7042/5/08Terpene Indole Alkaloidstryptophan

S. OConnorBiochemistry 7042/5/08Terpene Indole Alkaloidstryptophanhere for sarpagine

S. OConnorBiochemistry 7042/5/08Terpene Indole Alkaloidstryptophan