FCH 532 Lecture 31

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FCH 532 Lecture 31

• Chapter 28 Nucleotide metabolism
• Quiz on Mon (4/16) IMP synthesis-Purine
  synthesis
• Quiz on Wed(4/18) Pyrimidine biosynthesis/regulat
  ion
• Quiz on Friday(4/20) Ribonucleotide reductase
  mechanism
• Friday (4/20) extra credit seminar, Dr. Jimmy
  Hougland,
• 145 Baker, 3-4PM.
• ACS exam has been moved to Monday (4/30)
• Quiz on Final is scheduled for May 4,
  1245PM-245PM, in 111 Marshall

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Page 1080
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Proposed mechanism for rNDP reductase
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Proposed reaction mechanism for ribonucleotide
reductase

• Free radical abstracts H from C3
• Acid-catalyzed cleavage of the C2-OH bond
• Radical mediates stabilizationof the C2 cation
  (unshared electron pair)
• Radical-cation intermediate is reduced by
  redox-active sulhydryl pair-deoxynucleotide
  radical
• 3 radical reabstracts the H atom from the
  protein to restore the enzyme to the radical
  state.

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Catabolism of purines

• All pathways lead to formation of uric acid.
• Intermediates could be intercepted into salvage
  pathways.
• 1st reaction is the nucleotidase and second is
  catalyzed by purine nucleoside phosphorylase
  (PNP)
• Ribose-1-phosphate is isomerized by
  phosphoribomutase to ribose-5-phosphate
  (precursor to PRPP).
• Purine nucleoside Pi
  Purine base ribose-1-P
• Adenosine and deoxyadenosine are not degraded by
  PNP but are deaminated by adenosine deaminase
  (ADA) and AMP deaminase in mammals

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Figure 28-23 Major pathways of purine catabolism
in animals.
ADA
Genetic defects in ADA kill lymphocytes and
result in severe combined immunodeficiencey
disese (SCID). No ADA results in high levels of
dATP that inhibit ribonucleotide reductase-no
other dNTPs
Page 1093
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Figure 28-24a Structure and mechanism of
adenosine deaminase. (a) A ribbon diagram of
murine adenosine deaminase in complex with its
transition state analog HDPR.
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Figure 28-24b (b) The proposed catalytic
mechanism of adenosine deaminase.

• Zn2 polarized H2O molecule nucleophilically
  attacks C6 of the adenosine. His is general base
  catalyst, Glu is general acid, and Asp orients
  water.
• Results in tetrahedral intermediate which
  decomposes by elimination of ammonia.
• Product is inosine in enol form (assumes dominant
  keto form upon release from enzyme).

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Purine nucleotide cycle

• Deamination of AMP to IMP combined with synthesis
  of AMP from IMP results in deaminating Asp to
  yield fumarate.
• Important role in skeletal muscle-increased
  activity requires increased activity in the
  citric acid cycle.
• Muscle replenishes citric acid cycle
  intermediates through the purine nucleotide
  cycle.

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Figure 28-25 The purine nucleotide cycle.
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Xanthine oxidase

• Xanthine oxidse (XO) converts hypoxanthine to
  xanthine, and xanthine to uric acid.
• In mammals, found in the liver and small
  intestine mucosa
• XO is a homodimer with FAD, two 2Fe-2S clusters
  and a molybdopterin complex (Mo-pt) that cycles
  between Mol (VI) and Mol (IV) oxidation states.
• Final electron acceptor is O2 which is converted
  to H2O2
• XO is cleaved into 3 segments. The uncleaved
  enzyme is known as xanthine dehydrogenase (uses
  NAD as an electron acceptor where XO does not).
• XO hydroxylates hypoxanthine at its C2 position
  and xanthine at the C8 positon to produce uric
  acid in the enol form.

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Figure 28-26a X-Ray structure of xanthine
oxidase from cows milk in complex with salicylic
acid.
N-terminal domain is cyan Central domain is
gold C-terminal domain is lavender
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Mechanism for XO

• Reaction initiated by attack of enzyme
  nucleophile on the C8 position of xanthine.
• The C8-H atom is eliminated as a hydride ion that
  combines with Mo (VI) complex, reducing it to Mo
  (IV).
• Water displaces the enzyme nucleophile producing
  uric acid.

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Figure 28-27 Mechanism of xanthine oxidase.
Page 1096
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Figure 28-23 Major pathways of purine catabolism
in animals.
ADA
Genetic defects in ADA kill lymphocytes and
result in severe combined immunodeficiencey
disese (SCID). No ADA results in high levels of
dATP that inhibit ribonucleotide reductase-no
other dNTPs
Page 1093
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Purine degredation in other animals
Primates, birds, reptiles, insects-final
degradation product id uric acid which is
excreted in urine. Goal is the conservation of
water.
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Figure 28-29 The Gout, a cartoon by James Gilroy
(1799).
Page 1097
Gout is a disease characterized by elevated
levels of uric acid in body fluids. Caused by
deposition of nearly insoluble crystals of sodium
urate or uric acid.
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Clinical disorders of purine metabolism
Excessive accumulation of uric acid Gout
The three defects shown each result in elevated
de novo purine biosynthesis
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Common treatment for gout allopurinol
Allopurinol is an analogue of hypoxanthine that
strongly inhibits xanthine oxidase. Xanthine
and hypoxanthine, which are soluble, are
accumulated and excreted.
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Catabolism of pyrimidines

• Animal cells degrade pyrimidines to their
  component bases.
• Happen through dephosphorylation, deamination,
  and glycosidic bond cleavage.
• Uracil and thymine broken down by reduction (vs.
  oxidation in purine catabolism).

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Biosynthesis of of NAD and NADP Produced from
vitamin precursors Nicotinate and Nicotinamide
and from quinolinate, a Trp degradation product
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Biosynthesis of FMN and FAD from riboflavin
FAD is synthesized from riboflavin in a
two-reaction pathway. Flavokinase phosphorylates
the 5OH group to give FMN FAD pyrophosphorylase
catalyzes the next step (coupling of FMN to ADP).
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Biosynthesis of CoA from pantothenate
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