Purine And Pyrimidine Biosynthesis, Nucleotide Metabolism And Regulation

Front 1

in general what are the 2 types of pathways that lead to nucleotides

Back 1

-de novo from AA, ribose 5-phosphate, CO2 and NH3; requires ATP
-salvage pathway recycles free bases and nucleosides released from nucleic acid breakdown via activated ribose (5-phosphoribosyl-1-pyrophosphate from ribose5-phosphate in pentose phosphate pathway) PRPP + base to yield nucleotide

Front 2

describe the de novo synthesis of purines

Back 2

-parent nucelotides are AMP and GMP
-purine ring is assembled on ribose phosphate
ribose-5-phosphate + ATP -> 5PRPP + AMP via PRPP synthetase
-PRPP is limiting substance; ADP and GDP are neg effectors of enzyme
-1st committed step: amino group from glutmatine is attached at C1 of PRPP by glutamine phosphoribosyl amidotransferase yileding 5-phosphoribosylamine which is highly usable with half life of 30s; pruine ring build on this structure
-assembly of ring yields IMP: 2 steps require THF cofactor; aspartate donates amino group in formation of amide bond followed by elimination of carbon skeleton of aspartate (as fumarate) (analogous to role in urea cycle)
-Pemetrexed was approved for cancer treatment by inhibiting folate enzymes dihydrofolate reductase and thymidylate synthase

Front 3

describe enzymes of iMP synthesis and first intermediate to have a complete purine ring

Back 3

-large multi-enzyme complexes; channeling of intermediates from one enzyme to next in these complexes imp for unstable intermediates
-first intermediate with complete ring is insinuate or IMP

Front 4

describe IMP to adneylate

Back 4

-requires insertion of amino group derived from aspartate in 2 rxns
-GTP is used in synthesizing adenylosuccinate; guanylate formed by NAD+ oxidation of inosinate at C2 followed by addition of amino from glutamine and ATP->AMP yields GMP

Front 5

describe biosynthesis of pyrimidines

Back 5

-not synthesized as nucleotide derivatives
-pyrimidine ring is completed before ribose-5-phosphate is attached
-only carbomyl phosphate and aspartate are only precursors that contribute atoms to ring
1. bicarbonate + NH4 and glutamine + 2ATP yiled carbamoyl phosphate which donates C2 and N3 to ring' aspartate donates other atoms
-carbamoyl phosphate synthesized by carbamoyl phosphate synthetase 2: 2 chains- one for glutamine hydrolysis to generate ammonia, larger 1 ATP site for bicarbonate to carboxyphosphate which reacts with ammonia to generate carbamic acid, other ATP site for carbamic acid to carbamoyl phosphate

Front 6

describe CAD

Back 6

CPS2, aspartate transcarbamoylase and dihydroorotase is multifunctional enzyme- carbamoyl phosphate to carbamoylaspartate to dihydroorotate to orotate
-immunosuppressive drug leflunomide treats rheumatoid arthritis inhibits at dihydroorotate dehydrogenase (dihydroorotate to orotate)

Front 7

describe orotate to end of pyrimidine synthesis

Back 7

-orotate couples to ribose in PRPP to yield orotidylate, a pyrimidine nucleotide, driven by hydrolysis of PPi via orotate phosphoribosyltransferase
-orotidylate is then decarboxylated to form uridylate UMP via orotidylate decarboxylase
-orotate phosphoribosyltransferase and orotidylate decarboxylse found on bifunctional protein UMP synthase

Front 8

dsescribe hereditary orotic aciduria

Back 8

-rare genetic disorder
-severe anemia, growth retardation and high levels of orotic acid excretion
-defect in one or both of activities of UMP synthase
-treatment high doses or uridine which leads to dec formation of orotic acid and to reversal of hematologic problem
-uridine is taken up by cells and converted to UMP, UDP then UTP which inhibits CPS2
-UTP is also substrate for CTP synthesis

Front 9

describe synthesis of cytidine

Back 9

-synthesized from uracil base of UMP but UMP is converted to UTP before synthesis can take place
-nucleoside monophosphate kinases utilize ATP to convert mono to diphosphates
-nucleoside diphosphates and triphosphates are interconverted by nucleoside diphosphate kinase by nucleoside diphosphate kinase (broader specificity than monophosphate kinases)
-UTP can be transformed into CTP by replacement of carbonyl group by amino group; requires ATP and uses glutamine for amino- O4 atom is phosphorylated to form reactive intermediate and phosphate displaced from by ammonia

Front 10

describe formation of thymidylate

Back 10

-formed by methylation of deoxyuridylate
-UDP to dUDP via ribonucleotide reductase
-dUDP to dUTP via nuceloside diphosphate kinase
-dUTP to dUMP dUTPase via N5N10methylenetetrahydrofolate donating C and oxidizes THF to dihydrofolate which is reduced to THF by dihydrofolate reductase
-dUMP to dTMP via thymidylate synthase
-note CDP to dCDP to dCTP to dUTP
-Flurouracil is a potent anticancer drug; F-dUMP irreversibly inhibits thymidylate snythase via suicide inhibition

Front 11

describe salvage pathway in nucleotide biosynthesis

Back 11

-resynthesis of nucleotides from bases via phosphoribosyltransferases; PRPP to nucleoside 5-monophosphate
-Purine- free purines released in nucleic acid turnover (particularly mRNA) in form of adenine, guanine and hypoxanthine (base in IMP); adenine phosphoribosyltransferase mediate AMP formation and hypoxanthine-guanine phosphoribosyltransferase (HGPRTase) can act on guanine or hypoxanthine to form GMP or IMP
-Pyrimidine- conversion of nucleotides by pyrimidine phosphoribosyltransferases- enzyme from human erythrocytes utilizes orotate, uracil and thymine as substrate but no cytosine

Front 12

describe Lesch-Nyhan Syndrome

Back 12

-consequence of mutations in salvage pathway
-hallmark symptom- compulsive self injury
-nearly total absence of hypoxanthine-guanine phosphoribosyltransferase HGPRTase
-hyperuricemia-can lead to kidney stones early in life and gout later and neurological problems such as spasticity, mental retardation and self mutilation; at age 2 or 3 children begin to bite fingers and will chew off in unrestrained
-HGPRTase gene on X chromosome so sex-linked recessive disorder limited to males, if gene is less than 2% normal mental retardation is present and less than .2% self mutilation is present
-patients usually die from kidney failure

Front 13

describe formation of NAD, FAD, CoA

Back 13

-formed from ATP
-nicotinate (niacin or vitamin B3) is derived from tryptophan; humans can synthesize required amt of nicotinate if supply of tryptophan in diet is adequate (dietary deficiency of both can lead to pellagra- dermatitis, diarrhea, and dementia; endocrine tumor that consumes large amts of tryptophan synthesizes serotonin can lead to pellagra-like symptoms)
-NICOTINATE + PRPP -> nicotinate ribonucleotide + ATP + glutamine yields NAD+
-riboflavin + ADP -> riboflavin 5-phosphate (FMN) + ADP

Front 14

describe synthesis of deoxyribonucleotides

Back 14

-ribonucleotide diphosphate + NADPH to deoxyribonucleotide diphosphate + NADP+ + H2O via ribonucleotide reductase

Front 15

describe regulation of nucleotide biosynthesis

Back 15

-feedback inhibition
-purine- balance between AMP and GMP production; committed step is conversion of PRPP to phosphoribosylamine by glutamine phosphoribosyl amidotransferase; salvage pathway- PRPP is consumed, dec rate of formation of 5phosphoribosylamine and AMP and GMP inhibit; insinuate is branch point of AMP and GMP synthesis, AMP and GMP inhibitto adenylosuccinate and xanthylate after IMP
-reduction of ribonucleotides to deoxyribonucleotides is controlled by allosteric interactions' ribonucleotide reductase activity is diminished by binding of dATP which signals abundance of deoxyribonucleotides and binding of ATP reverses inhibition; binding of dATP or ATP enhances reduction of UDP and CDP
-pyrimidine regulation- at CPS2 inhibited by UTP, activated by PRPP
-CPS2 is only source of carbamoyl phosphate in extrahepatic tissues except in stressed conditions (excess ammonia) CPS1 generate carbamoyl phosphate in mitochondria which ends up in cytosol and serves as substrate for pyrimidine nucleotide synthesis, detoxifying ammonia leading to elevated excretion of orotic acid
-UMP does not inhibit CPS2 but it competes with OMP to inhibit OMP decarboxylase

Front 16

describe degradation of nucleotides

Back 16

-nucleic acids via nucleases to nucleotides
-nucleotides hydrolytically degraded to nucleosides by nucleotidases
-phosphoryltic cleavage of nucleosides to free bases and ribose 1-phosphate or deoxyribose-1phosphate by nucleoside phosphorylases
-ribose1-phosphate isomerized to ribose 5-phosphate, substrate in synthesis of PRPP

-purine bases to xanthine to uric acid to urate (excreted)
--eg AMP to adenosine to hypoxanthine through deamination by adenosine deaminase to xanthine via xanthine oxidase to uric acid to urate (note O2 reduced to H2O2)
-pyrimidine bases- thymine to B-aminoisobutyric acid; uracil to B-alanine (ring); NH4 production and urea synthesis
-average serum level of urate is close to solubility limit; urate is scavenger for ROS (as effective as ascorbate (vit C))

Front 17

describe immunodeficiency diseases associate with adenosine deaminase (ADA)

Back 17

-severe combined immunodeficiency involving T-cell functions with sparing of effects on B-cell function
-intracellular concentrations of dATP and S-adenosylhomocystein are greatly inc
-treatment includes blood transfusions, bone marrow transplantation, enzyme replacement therapy with ADA-PEG and gene therapy

Front 18

describe gout

Back 18

-high serum levels of urate induce gout- salts of urate and uric acid crystallize and damage joints and kidneys
-plasma urate greater than 7 mg/dL
-in joints causes inflammation arthritis and pain
-treatment allopurinol, inhibitor of xanthine oxidase; dec uric acid formation and synthesis of purine nucleotides therefore cancer treatment
-hyperuricemia from overproduction of purine nucleotides can be distinguished from hyperuricemia from kidney disease or cell death by feeind N15 glycine to pt who is overproducer results in uric acid excreted in urine with N15 at N7 of uric acid while pt not overproducer no N15 in uric acid
-those predisposed to gout are advised to reduce intake of purine-rich foods including red meat, game, seafood and alcoholic drinks especially beer

Front 19

describe biochemical defects resulting in inc de novo purine nucleotide synthesis

Back 19

-inc PRPP synthetase- positive effector of glutamine PRPP amidotransferase
-dec HGPRTase activity- dec salvage of hypoxanthine/guanine inc PRPP and IMP GMP not negative effectors
--in overproducers allopurinol treatment inhibits xanthine oxidase and purine bases are then salvaged via HGPRTase to form IMP and XMP

Front 20

describe clinical correlation of thymine degradation and cancer pts

Back 20

-inc levels of B-aminoisobutyrate are excreted in cancer patients undergoing chemo or radiation; also inc concentrations of uric acid being excreted and in serum
-due to destruction of tumor cells that in turn release degraded nucleic acids and nucleotides that are further metabolized to B-aminoisobutyrate and uric acid

Front 21

describe infantile autsim and uric acid excretion

Back 21

-subclass of children with infantile autism were shown to excrete uric acid; represents 20% of autistic population
-inc synthesis of purine nucleotides 4 fold observed
-molecular basis unknown
-serum concentrations of uric acid were in normal range