Amino Acid and Nucleotide Biosynthesis Lecture Notes PDF

Amino acid and nucleotide biosynthesis Marc A. Ilies, Ph. D. Lehninger - Chapter 22 [email protected]; lab 517, office 517A (Tu, Fr 3-5) For questions, comments please use the discussion tool in Canvas...

Amino acid and nucleotide biosynthesis Marc A. Ilies, Ph. D. Lehninger - Chapter 22 [email protected]; lab 517, office 517A (Tu, Fr 3-5) For questions, comments please use the discussion tool in Canvas ©MAIlies 2024 1  Ammonia is incorporated into biomolecules through Glu and Gln Glutamine is made from Glu by glutamine synthetase in a two-step process. Phosphorylation of Glu creates a good leaving group that can be easily displaced by ammonia. Biosynthesis of amino acids and nucleotides― multiple transaminations Non-essential amino acids are made from ammonia and -keto acids, via transamination Transaminations use pyridoxal phosphate (PLP) PLP is active form of vitamin B6 Catalyzed by amidotransferases PLP has aldehyde group that forms Schiff base with Lys of aminotransferase  Amino acid synthesis overview Bacteria can synthesize all 20. Mammals require some in diet. Source of N is Glu or Gln Non-essential amino acids are synthesized from 7 intermediates of: – glycolysis – citric acid cycle – pentose phosphate pathway  Amino Acid Biosynthetic Families, Grouped by TABLE 22-1 Metabolic Precursor α-Ketoglutarate Pyruvate Glutamate Alanine Glutamine Valinea Proline Leucinea Arginine Isoleucinea 3-Phosphoglycerate Phosphoenolpyruvate and Serine erythrose 4-phosphate Glycine Tryptophana Cysteine Phenylalaninea Tyrosineb Oxaloacetate Ribose 5-phosphate Aspartate Histidinea Asparagine Methioninea Threoninea Lysinea a Essential amino acids in mammals. b Derived from phenylalanine in mammals. Biosynthesis of some special amino acids, biogenic amines, CNS-active compounds All are decarboxylated using PLP dependent enzymes. 6  Biosynthesis of porphyrins - starts from succinylCoA and glycine: Heme 7  Defects in heme biosynthesis generate porphyrias Most animals synthesize their own heme. Many drugs can Mutations or misregulaton of enzymes precipitate attacks of in the heme biosynthesis pathway lead to porphyrias. – Precursors accumulate in red blood cells, body fluids, and liver. Accumulation of precursor uroporphyrinogen – Urine becomes discolored (pink to dark purplish depending on light, heat exposure). – Teeth may show red fluorescence under UV light. – Skin is sensitive to UV light. – There is a craving for heme. Explored as possible biochemical basis for vampire myths 8  Heme is the source of bile pigments Heme Heme from processing of old erythrocytes is Degradation degraded to bilirubin in two steps: biliverdin 1. Heme oxygenase linearizes heme to 15Z create biliverdin, a green compound 4Z (seen in a bruise). 2. Biliverdin reductase converts biliverdin to bilirubin, a yellow compound that travels bound to serum albumin in the bloodstream. major pigment of urine Linear Tetrapyrrole (degradation to urobilin) further degraded by intestinal microbiota to stercobilin UDP-glucuronosyl transferase Jaundice is caused by accumulation of bilirubin Excreted via glucuronidation often from liver disease (liver cancer, hepatitis), blocked bile secretion (gallstones, pancreatic cancer) or lack of UDPGT (Crigler-Najjar) Infant jaundice phototherapy: 4Z,15Z (cis-cis) is toxic; blue light converts to 4Z,15E and intramolecular cyclization to 9 soluble non-toxic product(s)  Biosynthesis of purines PRPP De-novo purine synthesis starts with a sugar (PRPP) and units are added to form a complete nucleotide First enzyme (glutamine-PRPP amidotransferase) is the rate-limiting step; inhibited by anticancer drugs SH 6-mercaptopurine SH H and 6-thioguanine) H N N N N N 10 N N H2N N  Biosynthesis of pyrimidines ► Unlike purine synthesis, pyrimidine synthesis proceeds by first making the pyrimidine ring (in the form of orotate) and then attaching it to ribose 5- phosphate. ► Aspartate and carbamoyl phosphate provide the atoms for the ring structure: HN Carbamoyl Aspartic acid phosphate N H ► UMP is phosphorylated to UTP. ► After formation of UTP, amination can convert UTP to CTP. 11  Ribonucleotides are precursors to Deoxyribonucleotides 2’C-OH bond is directly reduced to 2’-H bond through the action of ribonucleotide reductase, using NADPH as reducing agent, via glutathione and FAD: Reduction performed on NDPs!  dTMP is made from dUMP ► reaction dUMP  dTMP catalyzed by thymidylate synthase, which adds a methyl group from tetrahydrofolate Thymidylate synthase is the THF DHF target for anticancer drugs 5- fluorouracil (5FU), floxuridine, capecitabine (irreversible inhibitors) Since both purines and pyrimidines require THF, made from DHF via dihydrofolate reductase DHFR, another anticancer strategy is to block DHFR with anticancer drug methotrexate (very toxic!)  Pyrimidines and purine degradation - start with nucleases to degrade DNA and RNA to individual nucleotides - resulted nucleotides can be re-cycled to avoid de-novo synthesis (salvage pathway); excess is catabolized Salvage Pathway converts purines & their nucleosides to mononucleotides Hypoxanthine Guanine Hypoxanthine-Guanine PRPP PRPP Phosporibosyl Transferase PPi (HGPRT) PPi IMP GMP Lesch-Nyhan Disease: Total lack of HGPRT Symptoms: severe mental retardation and deposits of orange sand like crystals in the diapers of infants afflicted with this condition. 14  Purine catabolism 1. Dephosphorylation (via 5’-nucleotidase) 2. Deamination and hydrolysis of ribose lead to production of xanthine. 3. Hypoxanthine and xanthine are then oxidized into uric acid by xanthine oxidase (XO). 4. Uric acid is excreted (in humans) and it can be converted to allantoin (in most mammals) and excreted 15  Excess uric acid causes Gout Painful joints (often in toes) due to deposits of sodium urate crystals Primarily affects males May involve overproduction or genetic under-excretion of uric acid and/or may involve overconsumption of fructose; also due to HGPRT deficiency from the salvage pathway Treated with avoidance of purine-rich foods (seafood, liver) or avoidance of fructose Also treated with xanthine oxidase inhibitors allopurinol (Zyloprim) or febuxostat (Uloric): Febuxostat Also treated by (Uloric®) administration of porcine uric acid oxidase (uricase) (competitive inhibitor, tightly bound to XO) (Krystexxa)  Catabolism of Pyrimidines Leads to NH4+ and urea Can produce intermediates of CAC (e.g. thymine is degraded to succinyl- CoA): succinylCoA  Goals and Objectives Upon completion of this lecture at minimum you should be able to answer the following: ►What is the main reaction used for amino acid biosynthesis, which class of enzymes is catalyzing it, what coenzymes it uses? What is the primary source of ammonia in amino acid biosynthesis? ► Which are the main metabolic precursors of amino acid biosynthesis and which amino acids can be biosynthesized from each precursor? ► How are CNS active compounds DOPA, GABA, and the biogenic amines dopamine, epinephrine, norepinephrine, serotonin and histamine biosynthesized? ► Which starting materials and intermediates are involved in the synthesis of porphyrins and heme and what diseases can be observed at this level? ►How is heme processed (catabolized), which intermediates are generated, what diseases can be observed (with symptoms) and how they can be treated? ► Which are the particularities of the biosynthesis of purines and pyrimidines and what drugs can be used to block it and at what level? Which is the source of each atom in the structure of purines and pyrimidines? ►Which are the particularities of purine and pyrimidines catabolism, which key enzymes are involved, what diseases are observed and how these diseases can be treated (drugs and their targets)? 18  Drugs and Diseases ► Diseases: Porphyrias; Crigler-Najjar; Liver cancer, hepatitis, pancreatic cancer, gallstones: Jaundice; Infant jaundice; Cancer; Gout ► Drugs and supplements: essential amino acids, pyridoxal phosphate (PLP, B6), d6-mercaptopurine, 6-thioguanine, 5-fluorouracil (5FU), floxuridine, capecitabine, methotrexate, allopurinol (Zyloprim), febuxostat (Uloric), porcine uric acid oxidase (uricase, Krystexxa) 19