KelleyLect4_2009 - Nucleotide biosynthesis Nucleotide Roles of nucleotides 1)Precursors of nucleic acids synthesis 2)Universal energy currency

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Unformatted text preview: Nucleotide biosynthesis Nucleotide Roles of nucleotides: 1)Precursors of nucleic acids synthesis 2)Universal energy currency 3)Components of biosynthetic pathways 4)Signal transduction factors Nucleotide biosynthesis involves a Nucleotide complex series of reactions and transformations, but can be understood at a conceptual level given the common reaction mechanisms and precursors involved involved Purine biosynthesis Purine Pyrimidine Pyrimidine biosynthesis biosynthesis Nucleotide metabolism Nucleotide General concepts -Common intermediates/reactions in purine/pyrimidine syntheses - Transformation of intermediates into individual nucleotides - Chemical reactions used to degrade nts - Conversion of RNA nts into DNA nts Nucleotide nomenclature Nucleotide De novo nucleotide synthesis De 3 steps - Assembly/modification of purine/pyrimidine ring - Attachment of sugar - DNA: Conversion of ribose to deoxyribose Common intermediates in nt synthesis Common Common intermediates in nt synthesis Common Purine biosynthesis: Purine Synthesis of inosinate Part I Purine biosynthesis: Purine Synthesis of inosinate: Part II Displacement reactions in purine biosynthesis in Substrate activation by phosphorylation Substrate Functionalization of purines Functionalization Functionalization of purines Functionalization Pyrimidine biosynthesis Pyrimidine Pyrimidine biosynthesis Pyrimidine Carbamoyl phosphate synthetase Carbamoyl Multiple active sites - NH3 from glutamine - react bicarbonate with ATP - react carboxyphosphate with NH3 - react carbamic acid + ATP Substrate channeling in carbamoyl phosphate synthetase phosphate Substrates must Substrates traverse 80 Å traverse Pyrimidine biosynthesis + +H , O H2 - Attachment of ribose to orotate Attachment • Catalyzed by pyrimidine phosphoribosyltransferase • Mechanism unknown: may involve carbonium-type transition state at C1’ carbon followed by attack of orotate N. Decarboxylation of orotate Decarboxylation • catalyzed by orotidylate decarboxylase • CO2 evolved, resulting carbanion protonated by Lys residue Pyrimidine synthesis Pyrimidine Avoids accumulation of dUTP Conversion of UTP to CTP Conversion CTP synthetase catalyzes amination reaction Bacteria: NH3 is N donor Human: Glutamine is N donor Synthesis of dTMP Synthesis thymidylate synthase Synthesis of dTMP Synthesis Synthesis of dTMP Synthesis hydride transferred proton abstracted Drugs targeting dTMP synthesis Drugs Analogs of dihydrofolate Suicide inhibition of thymidylate synthase Suicide • Used clinically for treatment of skin, colon, breast, and stomach cancer • Drugs: Adrucil, Efudex, Fluoroplex Suicide inhibition of thymidylate synthase Suicide THF fU Purine biosynthesis Purine Pyrimidine Pyrimidine biosynthesis biosynthesis Ribonucleotide reductase Ribonucleotide Thiyl radical facilitates nt reduction Ribonucleotide reductase Mammals E. coli Yeast (most common) Thiyl radical Prokaryotes Small subset of prokaryotes Anaerobic prokaryotes Ribonucleotide reductase Ribonucleotide Ribonucleotide reductase Ribonucleotide Ribonucleotide reductase Active site Cys e- Ribonucleotide reductase Step 1 Step Thiyl radical generated by long-range ET, loss of H+ Ribonucleotide reductase Step 2 Step C-3’ H atom abstracted → C-3’ radical generated Ribonucleotide reductase Step 3 Step Protonation of 2’-OH by nearby Cys-SH promotes loss of H2O Ribonucleotide reductase Step 4 Step Hydride ion transferred to C-2’ → disulfide formed → radical back to C-3’ Ribonucleotide reductase Step 5 Step Hydrogen atom abstracted by C3’ → reduced nucleotide produced Ribonucleotide reductase Step 6 Step Ribonucleotide reductase reduced oxidized Ribonucleotide reductase Ribonucleotide Elimination of dUTP by dUTPase dUTPase: dephosphorylates dUTP Elimination of dUTP by dUTPase dUTPase: dephosphorylates dUTP Elimination of dUTP by dUTPase Differentiation of U from T Purine catabolism Purine Mo(VI) Mo(IV) Xanthine oxidase oxidase Urate: product of purine catabolism catabolism Causative agent in gout Summary: Nucleotide biosynthesis - Purines and pyrimidines synthesized via sequential reactions of activated substrates - Similar amination reactions involved in generation of A, G, and C. - Enzymes involved in methylation of U to generate T are drug targets - Protein-based radicals drive conversion of ribo nts to deoxy nts by RNR ...
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This note was uploaded on 01/23/2012 for the course BCHM 311 taught by Professor Kelley during the Spring '09 term at University of Toronto- Toronto.

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