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Biosynthetic pathways by pooja

as per GTU sem -7 students
biosynthetic pathways - shikimik acid pathway

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Biosynthetic pathways by pooja

  1. 1. Prepared by Pooja H. Khanpara Asst. Professor Apip, jamnagar Biosynthetic Studies & Basic Metabolic Pathways
  2. 2. What is Biosynthesis? Biosynthesis is a process of forming larger organic compounds from small subunits within a living organism. Biosynthesis is mainly done by enzymes. Biosynthesis is also known as anabolism since simple compounds are joined together to form macromolecules by enzymes. As an example, photosynthesis occurs inside the chloroplast. The light energy is converted into chemical energy during photosynthesis.  The larger molecule glucose is biosynthesized from water and carbon dioxide by photosynthetic organisms.(ATP, Enzyme, Cofactors)
  3. 3. What is the Difference Between Synthesis and Biosynthesis? Synthesis vs. Biosynthesis Synthesis refers to the formation of macromolecules from small molecules artificially. Biosynthesis refers to the formation of larger organic compounds from small molecules within a living organism. Process Synthesis is artificial and chemical. Biosynthesis is biological and catalyzed by enzymes. Resulting Polymers Synthesis can result in polymers which are organic or non-organic. Biosynthesis is biological and catalyzed by enzymes. Occurrence Synthesis occurs outside living organisms. Biosynthesis occurs within a living organism.
  4. 4. Biosynthesis of Primary Metabolites  Living plants are solar-powered biochemical and biosynthetic laboratory which manufactures both primary and secondary metabolites from air, water, minerals and sunlight.  The primary metabolites like sugars, amino acids & fatty acids that are needed for general growth & physiological development of plant which distributed in nature & also utilized as food by man.  The secondary metabolites such as alkaloids, glycosides, Flavonoids, volatile oils etc are biosynthetically derived from primary metabolites.  Biosynthetic reactions are replica of common organic reactions like catalytic reactions, phosphorylation, hydride transfer, oxidation, elimination, acylation, alkylation, reduction, condensation, rearrangement etc.
  5. 5. Metabolism & Metabolic Pathways Cell Metabolism: Process by which living cell process nutrient molecule & living state. Metabolic Pathway: A complete set of chemical reactions that occur in living cells, allowing cells to grow and reproduce, maintain their structures, and respond to their environments. Living cell require energy for biosynthesis, transport of nutrient, motility and maintenance. Energy is obtained from the catabolism of carbon compounds (carbohydrate) Carbohydrates are synthesized from CO2 and H2O in the present of light by photosynthesis.
  6. 6. ~ produce energy to the cell ~ requires energy glucose to glycogen
  7. 7. Major Metabolic Pathways Cellular respiration: Glycolysis Anaerobic respiration Kreb’s cycle / Citric acid cycle Oxidative phosphorylation Creation of energetic compounds from non-living matter: Photosynthesis (plants, algae cynobacteria) Chemosynthesis (some bacteria) Other pathways occurring in (most or) all living cell: Fatty acid oxidation (β-oxidation) Gluconeogenesis HMG-CoA reductase pathway (isoprene prenylation) Pentose phosphate pathway (hexose monophosphate) Porphyrin synthesis (or heme synthesis) pathway Urea cycle
  8. 8. Metabolites Metabolites are the intermediates & products of metabolism. The term metabolite is usually restricted to small molecules. A primary metabolite is directly involved in the normal growth, development, and reproduction. A secondary metabolite is not directly involved in those processes, but usually has important ecological function.
  9. 9. Importance of photosynthesis in formation of primary metabolites Photosynthesis is the process where plants convert sunlight into energy, then store it as carbohydrates, sugars, such as glucose. Photosynthesis may be the most important process in ecosystems, both brings in energy needed within the ecosystem, and produce oxygen (O2) needed for cellular respiration, and the production of more ATP. Photosynthesis has three basic steps: 1. Energy is captured from the sunlight. 2. Light energy is converted into chemical energy in the form of ATP and NADPH. 3. Chemical energy is used to power the synthesis of organic molecules (e.g. carbohydrates) from carbon dioxide (CO2).
  10. 10. Photosynthesis H2O + light + ADP + P ---> O2 + ATP + e- After the above steps occur in photosystem II, the electron is finally sent to photosystem I, where the following happens. e- + NADP+ + H ---> NADPH Now there are two high energy molecules, fully charged and ready to be used. Plants make more energy that it needs immediately, so the NADPH and ATP are used to make glucose as follows: CO2 + ATP + NADPH ---> C6H12O6 This happens in Calvin cycle.
  11. 11. Calvin Cycle The Calvin cycle is the last step in photosynthesis. The purpose of the Calvin Cycle is to take the energy from photosystem I and fix carbon. Carbon fixation means building organic molecules by adding carbon onto a chain. The following formula summarizes the Calvin cycle. C5 + CO2 + ATP + NADPH → C6H12O6 Where C5 is a five carbon molecule, such as pyruvate, when is recycled as glucose is synthesized. The first step in the Calvin cycle is for the 3C5 to bind with 3CO2, producing a six 3-carbon organic molecules (6C3). Next, 6ATP and 6NADPH energizes the binding of a C3 to make a 6- carbon molecule (C6), glucose. The remaining 5C3 continues moving through the Calvin cycle, being turned back into the starter C5 organic molecule.
  12. 12. Calvin Cycle
  13. 13. Glycolysis (Embden-meyerhoff pathway) Glycolysis represents an anabolic pathway common in both aerobic and anaerobic organisms. Sugars and polysaccharides are transformed into glucose or one of its phosphorylated derivatives before being processed any further. In the course of degradation, ATP is produced. Pyruvate may be regarded as the preliminary final product of the degradation. Pyruvate is fed into the citric acid cycle via an intermediate product. This pathway produces energy in the form of ATP. The starting product glucose is completely oxydized to water and carbon dioxide.
  14. 14. Citric Acid Cycle (Kreb’s cycle) The citric acid cycle, is the common mode of oxidative degradation in eukaryotes and prokaryotes. It accounts for the major portion of carbohydrate, fatty acid and amino acid oxidation and produces at the same time a number of biosynthetic precursors. The GTP generated during the succinate thiokinase (succinyl- CoA synthetase) reaction is equivalent to a mole of ATP by virtue of the presence of nucleoside diphosphokinase. The 3 moles of NADH and 1 mole of FADH2 generated during each round of the cycle feed into the oxidative phosphorylation pathway. Each mole of NADH leads to 3 moles of ATP and each mole of FADH2 leads to 2 moles of ATP. Therefore, for each mole of pyruvate which enters the TCA cycle, 12 moles of ATP can be generated.
  15. 15. Carbohydrate Utilization Storage carbohydrate such as the starch of plants or the glycogen of animals is made available for energy production by a process. As a result of this, the energy-rich carbohydrate is eventually oxidized to CO2and H2O. During the process, the hydrogen atoms liberated are carried by coenzymes into the cytochrome system, in which energy is released in stages, with the possible formation of ATP and ADP and inorganic phosphate. Eventually the hydrogen combines with oxygen to form water. The overall reaction of glucose in terms of ADP and ATP is C6H12O6 + 6CO2 + 38 ADP + 38P (inorganic) → 6H2O + 6CO2 + 38 ATP
  16. 16. The primary and secondary metabolites derived from carbon metabolism
  17. 17. difference between Primary and secondary metabolites
  18. 18. Biosynthetic Pathway of Secondary Metabolites It involves 3 basic mechanism: 1. Shikimic acid Pathway 2. Acetate-mevalonate pathway 3. Acetate malonate pathway
  19. 19. Shikimic acid Commonly known as its anionic form shikimate, is a cyclohexene, a cyclitol and a cyclohexanecarboxylic acid. Its name comes from the Japanese flower shikimi the Japanese star anise, Illiciumanisatum), from which it was first isolated in 1885 by Johan Fredrik Eykman. The elucidation of its structure was made nearly 50 years later. Shikimic acid is also the glycoside part of some hydrolysable tannins.
  20. 20. Shikimic Acid Pathway The Shikimic acid pathway is a key intermediate from carbohydrate for the biosynthesis of C6-C3units (phenyl propane derivative). The Shikimic acid pathway converts simple carbohydrate precursors derived from glycolysis and the pentose phosphate pathway to the aromatic amino acids. The shikimate pathway is a 7 step metabolic route used by bacteria, fungi, Algae, parasites, and plants for the biosynthesis of aromatic amino acids (phenylalanine, tyrosine, and tryptophan). This pathway is not found in animals; therefore, phenylalanine and tryptophan represent essential amino acids that must be obtained from the animal's diet.  Animals can synthesize tyrosine from phenylalanine, and therefore is not an essential amino acid except for individuals unable to hydroxylate phenylalanine to tyrosine).
  21. 21. Phosphoenolpyruvate and erythrose-4-phosphate react to form 2-keto3- deoxy7phosphoglucoheptonic acid, in a reaction catalyzed by the enzyme DAHP synthase. 2-keto3-deoxy7phosphoglucoheptonic acid is then transformed to 3- dehydroquinate (DHQ), in a reaction catalyzed by DHQ synthase. Although this reaction requires nicotinamide adenine dinucleotide (NAD) as a cofactor, the enzymic mechanism regenerates it, resulting in the net use of no NAD.
  22. 22. DHQ is dehydrated to 3-dehydroshikimic acid by the enzyme 3-dehydroquinate dehydratase, which is reduced to shikimic acid by the enzyme shikimate dehydrogenase, which uses nicotinamide adenine dinucleotide phosphate (NADPH) as a cofactor. The next enzyme involved is shikimate kinase, an enzyme that catalyzes the ATPdependent phosphorylation of shikimate to form shikimate 3-phosphate. Shikimate 3-phosphate is then coupled with phosphoenol pyruvate to give 5- enolpyruvylshikimate-3-phosphate via the enzyme 5-enolpyruvylshikimate-3- phosphate (EPSP) synthase. Then 5-enolpyruvylshikimate-3-phosphate is transformed into chorismate by a chorismate synthase.
  23. 23. Prephenic acid is then synthesized by a Claisen rearrangement of chorismate by Chorismate mutase. Prephenate is oxidatively decarboxylated with retention of the hydroxyl group by Prephenate dehydrogenase to give phydroxyphenylpyruvate, which is transaminated using glutamate as the nitrogen source to give tyrosine and α- ketoglutarate.
  24. 24. Role of Shikimic Acid Pathway: Starting Point in The Biosynthesis of Some Phenolics Phenyl alanine and tyrosine are the precursors used in the biosynthesis of phenylpropanoids. The phenylpropanoids are then used to produce the flavonoids, coumarins, tannins and lignin. Gallic acid biosynthesis Gallic acid is formed from 3-dehydroshikimate by the action of the enzyme shikimate dehydrogenase to produce 3,5- didehydroshikimate. The latter compound spontaneously rearranges to gallic acid. Other compounds Shikimic acid is a precursor for: Indole, indole derivatives and aromatic amino acid tryptophan and tryptophan derivatives such as the psychedelic compound dimethyltryptamine. & many alkaloids and other aromatic metabolites.
  25. 25. Uses: In the pharmaceutical industry, shikimic acid from the Chinese star anise (Illicium verum) is used as a base material for production of oseltamivir (Tamiflu). Target for drugs Shikimate can be used to synthesize (6S)-6-Fluoroshikimic acid, an antibiotic which inhibits the aromatic biosynthetic pathway. Glyphosphate, the active ingredient in the herbicide Roundup, kills plants by interfering with the shikimate pathway in plants. More specifically, glyphosate inhibits the enzyme 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS). "Roundup Ready" genetically modified crops overcome that inhibition.
  26. 26. Atropine
  27. 27. 2nd method Take Powder drug, ext with 95% alcohol(soxhlet hot percolation) Ethanolic ext. concentrate it Add dil. HCl & filter it extracted with pet. Ether (to remove impurities) Aq. Sol. Make alkaline with NH3 Extracted with CHCl3 (3 times) Combine exts. Evaporate chloroform under vaccum Again ext. with dil. Oxalic Acid Finally get crystal of Atropine & Hyoscyamine
  28. 28. 1. Thalleioquin Test: sol. of chinchona drug when treated with Bromin & ammonia it produce emerald green color. 2.
  29. 29. Ext. with chloroform further purification make acidic (Codein) make alkaline (NH3) ppt(Morphine) sol. (Narceine)
  30. 30. Isolation of Reserpine Roots are powdered & moisten with 10 % NaHCo3 & ext. with benzene untill give positive reaction with HgI2 Conc. It & add ether & dil. HCl again conc. It & separate acid layer. Again washed with ether. Make it alkaline with NH3 Then ext. with CHCl3 The CHCl3 ext. washed with 10% Na2CO3 Dry it & purify it by using methanol Get pure crystal of Reserpine
  31. 31. Isolation of Ephedrine Powder drug , moist with Na2CO3 & ext. with Benzene Filter it Take residue ext. With Dil. HCl Make alkaline by using K2CO3 & add CHCl3 Add Na2SO4 in CHCl3 solution & dried it Take residue, treat with Oxalic Acid ,warm, filter & cool it Get Ephedrine oxalate crystal
  32. 32. Chemical Test: Ephedrine in H2O + dil. HCl treated with  CuSO4 + NaOH  violate color  add ether  purple color  Aq. Layer shows blue color
  33. 33. Podophyllum Indian Podophyllum Synonym – Indian May apple, Wild lemon, Duck’s Foot, Hog Apple Biological source – It consists of the dried rhizome and root of Podophyllum hexandrum or Podophyllum emodi Family – Berberidaceae Use: purgative, tmt of cancer (Ovarian cancer), antirhematic, insecticidal activity