![TLC of products of [ 3 H]Farnesol-bisTEAP reaction](https://image.slidesharecdn.com/LinkedInslideshow-123181481234-phpapp01/85/Isoprenoids-19-320.jpg)
Isoprenoids
- 1. Isoprenoids R. KennedyKeller, Ph.D. Professor, Molecular Medicine, USF Health President, Isoprenoids, LC Grand Prismatic Spring, Yellowstone Natl Park Jim Peaco, NPS
- 2. Isoprenoids: What arethey and who cares? Definition : any compound derived from isopentenyl pyrophosphate (IPP) (2-methyl-buta-1,3-diene) IPP
- 3.
- 4. Isoprenoids in animalsCholesterol – structural component of membranes, precursor to bile acids, steroid hormones, Vit D Dolichyl phosphate – carries carbohydrate residues in glycoprotein synthesis Coenzyme Q – carrier in the electron transport chain Farnesyl and Geranylgeranyl – hydrophobic anchors on C-termini of various proteins Fat-soluble vitamins (A, D, E, K) Juvenile hormone (in insects, responsible for development and mating) – insecticide target
- 5. Coenzyme Q (a.k.a. ubiquinone ) and dolichyl phosphate (Dol-P): two compounds synthesized from IPP
- 6. Isoprenoid compounds inplants
- 7. Isoprenoids in plantsSterols (structure) Chlorophyll (light absorption) Carotenoids (energy transfer and photoprotection) Absisic acid (hormone) Zeatin (cytokines) Gibberillins (growth regulator) Isoprene (gas)
- 8. Drugs : taxol(anticancer); artemesinin (antimalarial) Rubber (latex, gutta-percha) Perfumes (geraniol, etc.) Cleansing Agents (PineSol ® ) Isoprenoids in modern life
- 9. How is IPPbiosynthesized?
- 10. In animals, IPPis synthesized from mevalonate (MVA)
- 11. In bacteria, plastidsand some protozoa, IPP is synthesized via methylerythritol phosphate (MEP)
- 12. Beyond IPP: Synthesisof terpenes (C10), sesquiterpenes (C15), diterpenes (C20) and larger molecules
- 13. IPP reacts withGPP to make FPP. In animals, FPP can be metabolized to cholesterol, Dol-P, ubiquinone, GGPP, or farnesylated proteins.
- 14. Isoprenoid Research: How best to synthesize the isoprenoid pyrophosphate intermediates?
- 15. Synthesis of IsoprenoidPyrophosphates - “ Classical” method (Cramer, F.; Rittersdorf, W.; Bohm, M. Liebigs Ann. Chem. 1962 , 654 , 180 ) - Poulter method (Davisson, V.J.; Woodside, A.B.; Poulter, C.D. Methods Enzymol. 1985 , 110 , 130-144)
- 16. Keller & Thompsonmodification of the Cramer method
- 17. Mix alcohol, TEAP,trichloroacetonitrile in ACN. Incubate 37 o C, 15 min. Apply reaction to silica chromatography Collect product. Yield: 30%. Products: mono, di, triphosphates. Time required: 2 h. Keller & Thompson procedure
- 18. ROH R-O-P-O -- = O - O - ACN Proposed mechanism for the phosphorylation reaction using bis- triethylammonium phosphate R= isopentenol, dimetylallyl alcohol, geraniol, farnesol, geranylgeraniol n
- 19. TLC of productsof [ 3 H]Farnesol-bisTEAP reaction
- 20. Preparative TLC offarnesol- bis TEAP reaction products
- 21. Monitoring of flashchromatography column in isolation of FPP using the method of Keller and Thompson
- 22.
- 23. TLC of productsprepared by the method of Keller & Thompson J. Chromatography 645, 1993, 161 IPP GPP FPP GGPP FMP FPPP GPP FPP GGPP SILICA RP
- 24. HPLC (straight phase)of FPP ( left ) prepared by the procedure of Keller & Thompson J. Chromatography 645, 1993, 161
- 25. Capillary electrophoresis ofisoprenoid pyrophosphates prepared by the procedure of Keller & Thompson J. Chromatography 737, 1996 , 325
- 26. Capillary electrophoresis resolvesgeranyl phosphate (GMP), citronellyl phosphate (CMP), and neryl phosphate (NMP), all of which are easily resolved from farnesyl phosphate (FMP). J. Chromatography 737, 1996, 325
- 27. 31 P NMRof Farnesyl mono, di, and triphosphate prepared by the procedure of Keller and Thompson J. Chromatography 737, 1996, 325 FP FPP FPPP
- 28. Biological activities ofisoprenoid pyrophosphates prepared by the procedure of Keller and Thompson Enzyme assays: Protein farnesyl transferase FPP + Protein-C-AAX Protein-C-AAX + PP i
- 29. Conclusion The isoprenoidpyrophosphates, widely used in studies of isoprenoid metabolism, can be rapidly prepared from the starting alcohols using bis (triethylammonium) phosphate and trichloroacetonitrile, followed by silica chromatography.
- 30. Opened 2005 atUSF Connect:
- 31.
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Editor's Notes
- #2 Aerial view of Grand Prismatic Spring; Midway & Lower Geyser Basin; Jim Peaco; July 2001 Yellowstone National Park Image by NPS Photo The Grand Prismatic Spring of Yellowstone National Park showing steam rising from hot and sterile deep azure blue water in the center surrounded by huge mats of brilliant orange algae and bacteria. The color of which is due to the ratio of chlorophyll to carotenoid molecules produced by the organisms. During summertime the chlorophyll content of the organisms is low and thus the mats appear orange, red, or yellow. However during the winter, the mats are usually dark green, because sunlight is more scarce and the microbes produce more chlorophyll to compensate, thereby masking the carotenoid colors. It is 90 meters (300 feet) across and 50 meters (160 feet) deep. In the center of the pool the water is 87° C (188° F)—too hot to support life. In the cooler water along the edges of the pool, however, colonies of thermophilic (heat-loving) cyano-bacteria and algae thrive. Yellow, orange, and red pigments are produced by the bacteria as a natural sunscreen. As a result, the pool displays a spectrum of colors from the bright blue of the center to the orange, red, and brown algal mats along the edges.