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Nukleotidnedbrytning
LPG001
Martin Lidell

Lecture outline

  • Nucleotides – short repetition of structural parts
  • Functions of nucleotides
  • Degradation of nucleic acids from food sources
  • Degradation of purine nucleotides
  • Degradation of pyrimidine nucleotides
  • Two diseases related to purine nucleotides
    • Gout – a very common disease
    • Adenosine deaminase deficiency – a very rare disease

What is a nucleotide?

  • Nucleotide = Phosphate(s) + Pentose + Nitrogenous base
  • Nucleoside = Pentose + Nitrogenous base
  • Adenosine monophosphate
  • OH (in ribose) or H (in deoxyribose)

The nitrogenous bases – purines and pyrimidines

  • Five bases
  • PURINES: Purine, Adenine, Guanine
  • PYRIMIDINES: Pyrimidine, Cytosine, Uracil (in RNA), Thymine (in DNA)
  • Two rings; two purines
  • Three pyrimidines; pyramide from above; CUT

Functions of nucleotides – some examples

  • Building blocks for DNA and RNA (store and translate genetic information)
  • Building blocks for important biomolecules (e.g. Coenzyme A)
  • Signaling molecules (both extra- and intracellular) (e.g. cAMP, adenosine signaling – a nucleoside)
  • “Activators” of biomolecules used for biosynthesis
    • UDP-Glucose (activated form of glucose; glucose donor in glycogen synthesis)

Overview of nucleotide metabolism

  • Nucleotides

    • De novo synthesis
    • Salvage synthesis (synthesis from reused nitrogenous bases and sugars)
    • DNA and RNA synthesis
    • Conversion to other important biomolecules
    • Degradation

  • From degradation:

    • Nitrogenous bases →
      • Reuse for nucleotide synthesis (salvage)
      • Further degradation →
        • Purines → Urate + Urea
        • Pyrimidines → Urea + Energy or energy-rich molecules
    • Sugar moiety →
      • Reuse for nucleotide synthesis (salvage)
      • Energy source (ATP or energy-rich molecules)

Expensive with de novo synthesis of nucleotides – the salvage pathway is cheaper

  • De novo pathway vs salvage pathway
  • PRPP; 5-Phosphoribosyl 1-pyrophosphate

Degradation of nucleic acids from food sources

  • Degradation of dietary nucleic acids occur in the small intestine
  • Nucleases, secreted by the pancreas, hydrolyze RNA and DNA to oligonucleotides
  • Oligonucleotides are further hydrolyzed by pancreatic phosphodiesterases, producing mononucleotides
  • In the intestinal mucosal cells (intestinal epithelial cells), nucleotidases remove the phosphate groups, releasing nucleosides that are further degraded to free bases and sugars by nucleosidases
  • The liberated bases can potentially be used in salvage pathways for nucleotide synthesis (however, at least the purines appear to be degraded to a large extent already in the intestinal cells)

Nucleotide degradation pathways

  • Nucleotides → Degradation →
    • Nitrogenous bases →
      • Reuse for nucleotide synthesis (salvage)
      • Further degradation →
        • Purines → Urate + Urea
        • Pyrimidines → Urea + Energy or energy-rich molecules
    • Sugar moiety →
      • Reuse for nucleotide synthesis (salvage)
      • Energy source (ATP or energy-rich molecules)

Degradation of purine nucleotides – formation of uric acid

  • GMP → (via nucleotidases) → Guanosine
  • Nucleotidases convert the nucleotides into nucleosides
  • Guanosine → Guanine (and further degradation)
  • Pathway towards hypoxanthine, xanthine and uric acid

Adenosine deaminase – an important enzyme in the degradation of adenosine

  • Adenosine is deaminated to inosine by adenosine deaminase
  • Toxic ammonia converted into urea in the liver
  • Parallel pathway: GMP → Guanosine → Guanine, etc.

The sugar parts are removed by nucleoside phosphorylase

  • GMP → Guanosine
  • Sugar phosphates options:
    1. Reuse for nucleotide synthesis (convert to PRPP)
    2. Use for energy production or generation of energy-containing molecules
  • The sugar parts are removed by nucleoside phosphorylase

GMP and AMP degradation converge at the level of xanthine that is further metabolized to uric acid

  • Toxic ammonia converted into urea in the liver
  • Uric acid (urate) excreted in the urine

Degradation of pyrimidine nucleotides

  • Pathways for CMP, UMP and dTMP
  • Intermediates within brackets refer to metabolites from dTMP degradation

CMP and UMP degradation converge at the level of uridine

  • Nucleotidases convert the nucleotides into nucleosides
  • CMP, UMP → Uridine
  • Toxic ammonia converted into urea in the liver

The sugar part is removed by a phosphorylase to generate the free pyrimidine bases

  • Options for sugar phosphates:

    1. Reuse for nucleotide synthesis (convert to PRPP)
    2. Use for energy production or generation of energy-containing molecules

  • Options for free bases:

    1. Reuse for nucleotide synthesis
    2. Use for energy production (ATP) or generation of energy-containing molecules

Complete degradation of nitrogenous bases for ATP production or generation of energy-containing molecules

  • From CMP and UMP:
    • Acetyl CoA (Propionyl CoA) → ATP, fatty acids or ketone bodies
  • From dTMP:
    • Succinyl CoA (CAC intermediate) → ATP or glucose production
  • Enzymes/intermediates:
    • Methylmalonate semialdehyde dehydrogenase
    • Propionyl CoA carboxylase
    • Methylmalonyl CoA
    • Methylmalonyl CoA mutase
  • Toxic ammonia converted into urea in the liver

What happens with the sugar moiety produced during nucleotide degradation?

  • Ribose-1-phosphate ↔ (Phosphopentomutase) ↔ Ribose-5-phosphate
  • Deoxyribose-1-phosphate ↔ Deoxyribose-5-phosphate
  • Ribose-5-phosphate can enter the pentose phosphate pathway (transketolase and transaldolase)
  • Deoxyribose-5-phosphate → (Deoxyribose phosphate aldolase) → Glyceraldehyde-3-phosphate + Acetaldehyde
  • Fructose-6-phosphate + Glyceraldehyde-3-phosphate → glycolysis/gluconeogenesis connection
  • Acetyl CoA from acetaldehyde → ATP, fatty acids or ketone bodies
  • Can be reused for nucleotide synthesis (converted to PRPP)

Endproducts used for energy production or generation of energy-containing molecules:

  • Fructose-6-phosphate and glyceraldehyde-3-phosphate: ATP or glucose production
  • Acetyl CoA: ATP, fatty acids or ketone bodies

Full degradation of pyrimidines and purines

Pyrimidines

  • Generate ammonia (NH₃) that is converted into UREA by the liver and excreted in the urine
  • Metabolites that can be used for energy production (ATP) or converted into energy-containing molecules such as glucose (liver), fatty acids and ketone bodies

Purines

  • Primarily generate URIC ACID (urate) that is excreted in the urine
  • Some ammonia is also produced; converted into urea by the liver

Gikt – frĂ„n ”the disease of kings” till folksjukdom

  • Vid för höga uratnivĂ„er i blodet (>6–7 mg/dl) fĂ€lls urat ut som saltkristaller (ofta natriumurat)
  • Kristallerna lĂ€gger sig i leder, senor och omgivande vĂ€vnad (vanligast Ă€r stortĂ„ns grundled) och orsakar dĂ€r inflammation
  • Vanligaste artritsjukdomen (uppskattad förekomst i Sverige, 1–2 % av befolkningen)
  • De höga uratnivĂ„erna i blodet beror antingen pĂ„ ökad syntes eller pĂ„ minskad utsöndring av urat
  • Beror oftast pĂ„ livsstilsfaktorer, lĂ€kemedelsbehandling eller annan sjukdom

Preventiva ÄtgÀrder inkluderar bland annat:

  • Minskat intag av alkohol. Vid metabolism av etanol bildas laktat som kompetitivt hĂ€mmar utsöndring av urat i tubuli
  • Minskat intag av purinrika livsmedel (frĂ€mst inĂ€lvsmat, sardiner, ansjovis och musslor, men Ă€ven övrig fet fisk, skaldjur och kött)

LĂ€kemedelsbehandling av gikt – strategi 1

  • HĂ€mma bildningen av urat genom att hĂ€mma enzymet xantinoxidas som ansvarar för sista steget i nedbrytningen av puriner
  • Exempel pĂ„ lĂ€kemedelssubstanser som hĂ€mmar produktionen av urinsyra:
    • Allopurinol (hypoxantinanalog)
    • Febuxostat

LĂ€kemedelsbehandling av gikt – strategi 2

  • HĂ€mma reabsorptionen av urat frĂ„n urinen i njurtubuli genom att inhibera urattransportörer (dessa Ă„terför normalt en stor del av utsöndrat urat till blodet)
  • Ger sĂ€nkta uratnivĂ„er i blodet dĂ„ mer urat avgĂ„r med urinen
  • Exempel pĂ„ substans: Probenecid

SvÄr kombinerad immunbrist (SCID)

  • SCID (Severe Combined Immunodeficiency) – samlingsnamn pĂ„ ett flertal ovanliga sjukdomar som beror pĂ„ avsaknad av immunceller som T- och B-lymfocyter, vilket leder till ett defekt immunsystem
  • Utan behandling leder SCID till svĂ„r infektionsbenĂ€genhet och drabbade individer avlider ofta redan under det första levnadsĂ„ret
  • Adenosindeaminasbrist; mycket ovanlig form av SCID i Sverige
  • Autosomal recessiv nedĂ€rvning (mutationer i ADA-genen orsakar dysfunktionellt adenosindeaminas)
  • En nĂ€rmast total brist pĂ„ immuncellerna T- och B-lymfocyter ses vid adenosindeaminasbrist

SvĂ„r kombinerad immunbrist (SCID) – till följd av adenosindeaminasbrist

Möjlig koppling mellan enzymdefekt och avsaknad av immunceller:

  • Muterat adenosindeaminas som förlorat sin funktion → ansamling av deoxyadenosin som omvandlas till dATP → syntes av övriga deoxyribonukleotider hĂ€mmas (dATP hĂ€mmar ribonukleotidreduktas) → syntes, replikation och reparation av skadat DNA hĂ€mmas → pĂ„verkar framförallt snabbt prolifererande celler (celltyper med hög omsĂ€ttning) som dĂ„ genomgĂ„r apoptos (”programmerat sjĂ€lvmord”)
  • T- och B-lymfocyter under utveckling Ă€r mycket snabbt prolifererande celler och tros dĂ€rför pĂ„verkas i speciellt hög grad av tillstĂ„ndet

Behandling – gĂ„r ut pĂ„ att ge tillgĂ„ng till ”friskt enzym”:

  • Hematopoetisk stamcellstransplantation (benmĂ€rgstransplantation) frĂ„n frisk donator
  • Enzymsubstitutionsbehandling, dvs enzymet ges som lĂ€kemedel (PEG-konjugat ADA injiceras subkutant)
  • Genterapi; ”frisk ADA-gen” introduceras i individens egna hematopoetiska stamceller

Genterapi vid adenosindeaminasbrist

  • Har utförts pĂ„ ett fĂ„tal individer dĂ€r det inte varit möjligt att hitta lĂ€mplig donator

Översikt:

  • Virus, med en frisk kopia av ADA-genen tillverkas
  • De virus man anvĂ€nder saknar förmĂ„gan att ge upphov till sjukdom men har kvar egenskapen att bygga in nya gener i vĂ„r arvsmassa
  • Virusen infekterar sedan hematopoetiska stamceller isolerade frĂ„n den sjuka individens benmĂ€rg och för pĂ„ sĂ„ sĂ€tt in den friska genen i dessa celler
  • Cellerna ges tillbaka till den sjuka individen som dĂ€rmed har fĂ„tt ”friska stamceller” som kan bilda friska T-lymfocyter

Sammanfattning av nukleotidnedbrytning

  • Nukleotider har flera viktiga funktioner förutom att bilda nukleinsyrorna DNA och RNA

  • Fem kvĂ€vebaser:

    • TvĂ„ puriner; tvĂ„ ringar; GA
    • Tre pyrimidiner; pyramid frĂ„n ovan; CUT

  • De novo syntes av nukleotider Ă€r dyrt vilket gör att baserna och sockerenheterna Ă„tervinns i hög grad

Om fullstÀndig nedbrytning av nukleotider:

  • Sockerdelen kan anvĂ€ndas direkt som energikĂ€lla (ATP) eller omvandlas till energirika produkter

  • KvĂ€vebaserna:

    • Puriner: URINSYRA (URAT) + mindre mĂ€ngd urea
    • Pyrimidiner: UREA + energirika molekyler som kan anvĂ€ndas för direkt produktion av ATP eller omvandlas till energirika produkter

  • Defekter i nukleotidmetabolism kan orsaka sjukdom:

    • Gikt; mycket vanlig artritsjukdom; uratkristaller i leder pga höga uratnivĂ„er i blodet
    • Adenosindeaminasbrist (form av SCID); mycket ovanlig sjukdom; defekt adenosinnedbrytning orsakar nĂ€rmast total brist pĂ„ T- och B-lymfocyter; mycket infektionskĂ€nsliga

LĂ€sanvisningar

  • Detta förelĂ€sningsmaterial

  • Biochemistry, 10th ed, Berg et al.

    • 2023 W.H. Freeman, Macmillian Learning
    • Kapitel 26: sidorna 809–810

  • InstuderingsfrĂ„gor – finns upplagt pĂ„ Canvas

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