1. Nucleotides: Synthesis and Degradation

2. Roles of Nucleotides
Precursors to nucleic acids (genetic material and non-protein
enzymes).
Currency in energy metabolism (eg. ATP, GTP).
Carriers of activated metabolites for biosynthesis
(eg. CDP, UDP).
Structural moieties of coenzymes (eg. NAD, CoA).
Metabolic regulators and signal molecules (eg. cAMP,
cGMP, ppGpp).

3. Biosynthetic routes: De novo and salvage pathways
De novo pathways
Almost all cell types have the ability to synthesize purine and pyrimidine nucleotides from low molecular weight precursors in amounts sufficient for their own needs.
The de novo pathways are almost identical in all organisms.
Salvage pathways
Most organisms have the ability to synthesize nucleotides from nucleosides or bases that become available through the diet or from degredation of nucleic acids.
In animals, the extracellular hydrolysis of ingested nucleic acids represents the major route by which bases become available.

4. Reutilization and catabolism of purine and pyrimidine bases
blue-catabolism
red-salvage pathways
endonucleases:
pancreatic RNAse
pancreatic DNAse
phosphodiesterases:
usually non-specific

5. PRPP: a central metabolite in de novo and salvage pathways
PRPP synthetase
Enzyme inhinited by AMP, ADP, and GDP. In E. coli, expression is repressed
by PurR repressor bound to either guanine or hypoxanthine.
Roles of PRPP: his and trp biosynthesis, nucleobase salvage pathways, de novo synthesis of nucleotides

6. Purine Nucleotide Synthesis

7. Example of a salvage pathway: guanine phosphoribosyl transferase
In vivo, the reaction is driven to the right by the action of pyrophosphatase
Shown: HGPRT, cells also have a APRT.

8. De novo biosynthesis of purines: low molecular weight precursors of the purine ring atoms

9. Synthesis of IMP
The base in IMP is called
hypoxanthine
Note: purine ring built up at
nucleotide level.
precursors:
glutamine (twice)
glycine
N10-formyl-THF (twice)
HCO3
aspartate
In vertebrates, 2,3,5 catalyzed
by trifunctional enzyme,
6,7 catalyzed by bifunctional
enzyme.

10. Pathways from IMP to AMP and GMP
G-1: IMP dehydrogenase
G-2: XMP aminase
A-1: adenylosuccinate synthetase
A-2: adenylosuccinate lyase
Note: GTP used to make AMP, ATP used to make GMP.
Also, feedback inhibition by
AMP and GMP.

11. Pathways from AMP and GMP to ATP and GTP
Conversion to diphosphate involves specific kinases:
GMP + ATP < > GDP + ADP Guanylate kinase
AMP + ATP < > 2 ADP Adenylate kinase
Conversion to triphosphate by Nucleoside diphosphate kinase (NDK):
GDP + ATP <------> GTP + ADP DG0’= 0
ping pong reaction mechanism with phospho-his intermediate.
NDK also works with pyrimidine nucleotides and is driven by mass action.

12. Allosteric regulation of purine de novo synthesis

13. Purine degredation
AMP deamination in muscle, hydrolysis in other tissues.
Xanthine oxidase:contains FAD, molybdenum, and non-heme iron.
In primates, uric acid is the end product, which is excreted.

14. Purine degredation in
other animals

15. Clinical disorders of purine metabolism
Excessive accumulation of uric acid: Gout
The three defects shown each result in elevated de novo
purine biosynthesis

16. Allopurinol is an analogue of hypoxanthine that strongly inhibits
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.

17. Diseases of purine metabolism (continued)
Lesch-Nyhan Syndrome: Severe HGPRT deficiency
In addition to symptoms of gout, patients display severe behavioral disorders, learning disorder, aggressiveness and hostility, including self-directed. Patients must be restrained to prevent self-mutilation. Reason for the behavioral disorder is unknown.
X-linked trait (HGPRT gene is on X chromosome).
Severe combined immune deficiency (SCID): lack of adenosine deaminase (ADA).
Lack of ADA causes accumulation of deoxyadenosine. Immune cells, which have potent salvage pathways, accumulate dATP, which blocks production of other dNTPs by its action on ribonucleotide reductase. Immune cells can’t replicate their DNA, and thus can’t mount an immune response.

18. De novo pyrimidine biosynthesis
Pyrimidine ring is assembled as the free base, orotic acid, which is them converted to the nucleotide orotidine monophosphate (OMP).
The pathway is unbranched. UTP is a substrate for formation of CTP.

19. Pyrimidine Synthesis

20. De novo synthesis of pyrimidines
1: carbamyl phosphate
synthase
2: aspartate
transcarbamylase
3: dihydroorotase
4: dihydroorotate DH
5: orotate
phosphoribosyl
tranferase
6: orotidylate
decarboxylase
7: UMP kinase
8: NDK
9: CTP synthetase
CAD=1,2,3
5 +6=single protein

21. Regulation of
pyrimidine
de novo synthesis

22. Catabolism of
pyrimidines

23. Overview of dNTP biosynthesis
One enzyme, ribonucleotide reductase,
reduces all four ribonucleotides to their
deoxyribo derivitives.
A free radical mechanism is involved
in the ribonucleotide reductase
reaction.
There are three classes of ribonucleotide
reductase enzymes in nature:
Class I: tyrosine radical, uses NDP
Class II: adenosylcobalamin. uses NTPs
(cyanobacteria, some bacteria,
Euglena).
Class III: SAM and Fe-S to generate
radical, uses NTPs.
(anaerobes and fac. anaerobes).

24. Structure of rNDP reductase (E. coli, ClassI)

25. Proposed mechanism
for rNDP reductase

26. Proposed reaction mechanism for ribonucleotide reductase

27. Sources of reducing power for rNDP reductase

28. Biological activities of thioredoxin

29. Regulation of activities of mammalian rNDP reductase

30. Salvage and de novo pathways to thymine nucleotides

31. Substrate recvognition by dUTPase

32. Relationship between thymidylate synthase and enzymes of tetrahydrofolate metabolism

33. Catalytic mechanism of thymidylate synthase

34. Regeneration of N5, N10-methylenetetrahydrofolate

35. Biosynthesis of NAD+ and NADP+

36. Biosynthesis of CoA from pantothenate

37. Proposed reaction mechanism for FGAM synthetase

38. The transformylation reactions are catalyzed by a multiprotein complex
components of the complex:
GAR transformylase (3)
AICAR transformylase (9)
serine hydroxymethyl transferase, trifunctional formylmethenyl-methylene-THF synthase (activities shown with asterisk)

39. Proposed catalytic mechanism for OMP decarboxylase

40. Reactions catalyzed by eukaryotic dihydroorotate dehydrogenase

41. Nitrogenous Bases Planar, aromatic, and heterocyclic
Derived from purine or pyrimidine
Numbering of bases is “unprimed”

42. Nitrogenous Bases Pyrimidines Purines N1: Aspartate Amine
C2, C8: Formate
N3, N9: Glutamine
C4, C5, N7: Glycine
C6: Bicarbonate Ion

43. Nucleotide Metabolism
PURINE RIBONUCLEOTIDES: formed de novo
i.e., purines are not initially synthesized as free bases
First purine derivative formed is Inosine Mono-phosphate (IMP)
The purine base is hypoxanthine
AMP and GMP are formed from IMP

44. Purine Nucleotides
Get broken down into Uric Acid (a purine)

45. Purine Nucleotide Synthesis
ATP is involved in 6 steps and an additional ATP is needed to form the first molecule (R5P)
PRPP in the first step of Purine synthesis is also a precursor for Pyrimidine Synthesis, His and Trp synthesis
Role of ATP in first step is unique– group transfer rather than coupling
In second step, C1 notation changes from a to b (anomers specifying OH positioning on C1 with respect to C4 group)
In step 3, PPi is hydrolyzed to 2Pi (irreversible, “committing” step)

46. Coupling of Reactions
Hydrolyzing a phosphate from ATP is relatively easy
G°’= kJ/mol
If endergonic reaction released energy into cell as heat energy, wouldn’t be useful
Must be coupled to an exergonic reaction
When ATP is a reactant:
Part of the ATP can be transferred to an acceptor: Pi, PPi, adenyl, or adenosinyl group in transferase reaction OR
ATP hydrolysis can drive an otherwise unfavorable reaction
(synthetase; “energase”)

47. Purine Biosynthetic Pathway
Coupling of some reactions on pathway organizes and controls processing of substrates to products in each step
Increases overall rate of pathway and protects intermediates from degradation
In animals, IMP synthesis pathway is coupled:
Reactions 3, 4, 6
Reactions 7, 8
Reactions 10, 11

48. IMP Conversion to AMP

50. IMP Conversion to GMP

51. Regulatory Control of Purine Nucleotide Biosynthesis
GTP is involved in AMP synthesis and ATP is involved in GMP synthesis (reciprocal control of production)
PRPP is a biosynthetically “central” molecule (why?)
ADP/GDP levels – negative feedback on Ribose Phosphate Pyrophosphokinase
Amidophosphoribosyl transferase is activated by PRPP levels
APRT activity has negative feedback at two sites
ATP, ADP, AMP bound at one site
GTP,GDP AND GMP bound at the other site
Rate of AMP production increases with increasing concentrations of GTP; rate of GMP production increases with increasing concentrations of ATP

52. Regulatory Control of Purine Biosynthesis
At level of IMP production:
Independent control
Synergistic control
Feedforward activation by PRPP
Below level of IMP production
Reciprocal control
Total amounts of purine nucleotides controlled
Relative amounts of ATP, GTP controlled

53. Purine Catabolism and Salvage
All purine degradation in animals leads to uric acid
Ingested nucleic acids are degraded by pancreatic nucleases, and intestinal phosphodiesterases in the intestine
Group-specific nucleotidases and non-specific phosphatases degrade nucleotides into nucleosides
Direct absorption of nucleosides
Further degradation
Nucleoside + H2O  base + ribose (nucleosidase)
Nucleoside + Pi  base + r-1-phosphate (n. phosphorylase)
NOTE: MOST INGESTED NUCLEIC ACIDS ARE DEGRADED AND EXCRETED.

54. Intracellular Purine Catabolism
Nucleotides broken into nucleosides by action of 5’-nucleotidase (hydrolysis reactions)
Purine nucleoside phosphorylase (PNP)
Inosine  Hypoxanthine
Xanthosine  Xanthine
Guanosine  Guanine
Ribose-1-phosphate splits off
Can be isomerized to ribose-5-phosphate
Adenosine is deaminated to Inosine (ADA)

55. Intracellular Purine Catabolism
Xanthine is the point of convergence for the metabolism of the purine bases
Xanthine  Uric acid
Xanthine oxidase catalyzes two reactions
Purine ribonucleotide degradation pathway is same for purine deoxyribonucleotides

56. Adenosine Degradation

57. Xanthosine Degradation
Ribose sugar gets recycled (Ribose-1-Phosphate  R-5-P )
– can be incorporated into PRPP (efficiency)
Hypoxanthine is converted to Xanthine by Xanthine Oxidase
Guanine is converted to Xanthine by Guanine Deaminase
Xanthine gets converted to Uric Acid by Xanthine Oxidase

58. Xanthine Oxidase A homodimeric protein
Contains electron transfer proteins
FAD
Mo-pterin complex in +4 or +6 state
Two 2Fe-2S clusters
Transfers electrons to O2  H2O2
H2O2 is toxic
Disproportionated to H2O and O2 by catalase

59. THE PURINE NUCLEOTIDE CYCLE
AMP + H2O  IMP + NH4+ (AMP Deaminase)
IMP + Aspartate + GTP  AMP + Fumarate + GDP + Pi (Adenylosuccinate Synthetase)
COMBINE THE TWO REACTIONS:
Aspartate + H2O + GTP  Fumarate + GDP + Pi + NH4+
The overall result of combining reactions is deamination of Aspartate to Fumarate at the expense of a GTP

60. Purine Nucleotide Cycle
In-Class Question: Why is the purine nucleotide cycle important in muscle metabolism during a burst of activity?

61. Adenosine Deaminase CHIME Exercise: 2ADA
Enzyme catalyzing deamination of Adenosine to Inosine
a/b barrel domain structure
“TIM Barrel” – central barrel structure with 8 twisted parallel b-strands connected by a-helical loops
Active site is at bottom of funnel-shaped pocket formed by loops
Found in all glycolytic enzymes
Found in proteins that bind and transport metabolites

62. Uric Acid Excretion Humans – excreted into urine as insoluble crystals
Birds, terrestrial reptiles, some insects – excrete isoluble crystals in paste form (conserve water)
Others – further modification :
Uric Acid  Allantoin  Allantoic Acid  Urea  Ammonia

63. Purine Salvage Adenine + PRPP  AMP + PPi
Adenine phosphoribosyl transferase (APRT)
Adenine + PRPP  AMP + PPi
Hypoxanthine-Guanine phosphoribosyl transferase (HGPRT)
Hypoxanthine + PRPP  IMP + PPi
Guanine + PRPP  GMP + PPi
(NOTE: THESE ARE ALL REVERSIBLE REACTIONS)
AMP,IMP,GMP do not need to be resynthesized de novo !

64. A CASE STUDY : GOUT
A 45 YEAR OLD MAN AWOKE FROM SLEEP WITH A PAINFUL AND SWOLLEN RIGHT GREAT TOE. ON THE PREVIOUS NIGHT HE HAD EATEN A MEAL OF FRIED LIVER AND ONIONS, AFTER WHICH HE MET WITH HIS POKER GROUP AND DRANK A NUMBER OF BEERS.
HE SAW HIS DOCTOR THAT MORNING, “GOUTY ARTHRITIS” WAS DIAGNOSED, AND SOME TESTS WERE ORDERED. HIS SERUM URIC ACID LEVEL WAS ELEVATED AT 8.0 mg/dL (NL < 7.0 mg/dL).
THE MAN RECALLED THAT HIS FATHER AND HIS GRANDFATHER, BOTH OF WHOM WERE ALCOHOLICS, OFTEN COMPLAINED OF JOINT PAIN AND SWELLING IN THEIR FEET.

65. A CASE STUDY : GOUT
THE DOCTOR RECOMMENDED THAT THE MAN USE NSAIDS FOR PAIN AND SWELLING, INCREASE HIS FLUID INTAKE (BUT NOT WITH ALCOHOL) AND REST AND ELEVATE HIS FOOT. HE ALSO PRESCRIBED ALLOPURINOL.
A FEW DAYS LATER THE CONDITION HAD RESOLVED AND ALLOPURINOL HAD BEEN STOPPED. A REPEAT URIC ACID LEVEL WAS OBTAINED (7.1 mg/dL). THE DOCTOR GAVE THE MAN SOME ADVICE REGARDING LIFE STYLE CHANGES.

66. Gout Impaired excretion or overproduction of uric acid
Uric acid crystals precipitate into joints (Gouty Arthritis), kidneys, ureters (stones)
Lead impairs uric acid excretion – lead poisoning from pewter drinking goblets
Fall of Roman Empire?
Xanthine oxidase inhibitors inhibit production of uric acid, and treat gout
Allopurinol treatment – hypoxanthine analog that binds to Xanthine Oxidase to decrease uric acid production

67. ALLOPURINOL IS A XANTHINE OXIDASE INHIBITOR A SUBSTRATE ANALOG IS CONVERTED TO AN INHIBITOR, IN THIS CASE A “SUICIDE-INHIBITOR”

68. Lesch-Nyhan Syndrome A defect in production or activity of HGPRT
Causes increased level of Hypoxanthine and Guanine ( in degradation to uric acid)
Also,PRPP accumulates
stimulates production of purine nucleotides (and thereby increases their degradation)
Causes gout-like symptoms, but also neurological symptoms  spasticity, aggressiveness, self-mutilation
First neuropsychiatric abnormality that was attributed to a single enzyme

69. Purine Autism 25% of autistic patients may overproduce purines
To diagnose, must test urine over 24 hours
Biochemical findings from this test disappear in adolescence
Must obtain urine specimen in infancy, but it’s difficult to do!
Pink urine due to uric acid crystals may be seen in diapers

70. Pyrimidine Ribonucleotide Synthesis
Uridine Monophosphate (UMP) is synthesized first
CTP is synthesized from UMP
Pyrimidine ring synthesis completed first; then attached to ribose-5-phosphate
N1, C4, C5, C6 : Aspartate
C2 : HCO3-
N3 : Glutamine amide Nitrogen

71. UMP Synthesis Overview
2 ATPs needed: both used in first step
One transfers phosphate, the other is hydrolyzed to ADP and Pi
2 condensation rxns: form carbamoyl aspartate and dihydroorotate (intramolecular)
Dihydroorotate dehydrogenase is an intra-mitochondrial enzyme; oxidizing power comes from quinone reduction
Attachment of base to ribose ring is catalyzed by OPRT; PRPP provides ribose-5-P
PPi splits off PRPP – irreversible
Channeling: enzymes 1, 2, and 3 on same chain; 5 and 6 on same chain

72. Pyrimidine Synthesis

73. UMP  UTP and CTP
Nucleoside monophosphate kinase catalyzes transfer of Pi to UMP to form UDP; nucleoside diphosphate kinase catalyzes transfer of Pi from ATP to UDP to form UTP
CTP formed from UTP via CTP Synthetase driven by ATP hydrolysis
Glutamine provides amide nitrogen for C4

75. OMP DECARBOXYLASE : THE MOST CATALYTICALLY PROFICIENT ENZYME
FINAL REACTION OF PYRIMIDINE PATHWAY
ANOTHER MECHANISM FOR DECARBOXYLATION
A CARBANION INTERMEDIATE (UNSTABLE)
MUST BE STABILIZED
BUT NO COFACTORS ARE NEEDED!
SOME OF THE BINDING ENERGY BETWEEN OMP AND THE ACTIVE SITE IS USED TO STABILIZE THE TRANSITION STATE
“PREFERENTIAL TRANSITION STATE BINDING”

77. Regulatory Control of Pyrimidine Synthesis
Differs between bacteria and animals
Bacteria – regulation at ATCase rxn
Animals – regulation at carbamoyl phosphate synthetase II
UDP and UTP inhibit enzyme; ATP and PRPP activate it
UMP and CMP competitively inhibit OMP Decarboxylase
*Purine synthesis inhibited by ADP and GDP at ribose phosphate pyrophosphokinase step, controlling level of PRPP  also regulates pyrimidines

78. Orotic Aciduria
Caused by defect in protein chain with enzyme activities of last two steps of pyrimidine synthesis
Increased excretion of orotic acid in urine
Symptoms: retarded growth; severe anemia
Only known inherited defect in this pathway (all others would be lethal to fetus)
Treat with uridine/cytidine
IN-CLASS QUESTION: HOW DOES URIDINE AND CYTIDINE ADMINISTRATION WORK TO TREAT OROTICACIDURIA?

79. Degradation of Pyrimidines
CMP and UMP degraded to bases similarly to purines
Dephosphorylation
Deamination
Glycosidic bond cleavage
Uracil reduced in liver, forming b-alanine
Converted to malonyl-CoA  fatty acid synthesis for energy metabolism

80. Deoxyribonucleotide Formation
Purine/Pyrimidine degradation are the same for ribonucleotides and deoxyribonucleotides
Biosynthetic pathways are only for ribonucleotides
Deoxyribonucleotides are synthesized from corresponding ribonucleotides

81. DNA vs. RNA: REVIEW DNA composed of deoxyribonucleotides
Ribose sugar in DNA lacks hydroxyl group at 2’ Carbon
Uracil doesn’t (normally) appear in DNA
Thymine (5-methyluracil) appears instead

82. Formation of Deoxyribonucleotides
Reduction of 2’ carbon done via a free radical mechanism catalyzed by “Ribonucleotide Reductases”
E. coli RNR reduces ribonucleoside diphosphates (NDPs) to deoxyribonucleoside diphosphates (dNDPs)
Two subunits: R1 and R2
A Heterotetramer: (R1)2 and (R2)2 in vitro

83. RIBONUCLEOTIDE REDUCTASE
R1 SUBUNIT
Specificity Site
Hexamerization site
Activity Site
Five redox-active –SH groups from cysteines
R2 SUBUNIT
Tyr 122 radical
Binuclear Fe(III) complex

84. E. coli Ribonucleotide Reductase:
Chime Exercise
E. coli Ribonucleotide Reductase:
3R1R and 4R1R: R1 subunit
1RIB and 1AV8: R2 subunit

85. Ribonucleotide Reductase R2 Subunit
Fe prosthetic group– binuclear, with each Fe octahedrally coordinated
Fe’s are bridged by O-2 and carboxyl gp of Glu 115
Tyr 122 is close to the Fe(III) complex  stabilization of a tyrosyl free-radical
During the overall process, a pair of –SH groups provide the reducing equivalents
A protein disulfide group is formed
Gets reduced by two other sulfhydryl gps of Cys residues in R1

86. Mechanism of Ribonucleotide Reductase Reaction
Free Radical
Involvement of multiple –SH groups
RR is left with a disulfide group that must be reduced to return to the original enzyme

87. RIBONUCLEOTIDE REDUCTASE
ACTIVITY IS RESPONSIVE TO LEVEL OF CELLULAR NUCLEOTIDES:
ATP ACTIVATES REDUCTION OF
CDP
UDP
dTTP
INDUCES GDP REDUCTION
INHIBITS REDUCTION OF CDP. UDP
dATP INHIBITS REDUCTION OF ALL NUCLEOTIDES
dGTP
STIMULATES ADP REDUCTION
INHIBITS CDP,UDP,GDP REDUCTION

88. RIBONUCLEOTIDE REDUCTASE
CATALYTIC ACTIVITY VARIES WITH STATE OF OLIGOMERIZATION:
WHEN ATP, dATP, dGTP, dTTP BIND TO SPECIFICITY SITE OF R1 (CATALYTICALLY INACTIVE MONOMER)
 CATALYTICALLY ACTIVE (R1)2
WHEN dATP OR ATP BIND TO ACTIVITY SITE OF DIMERS
 TETRAMER FORMATION
(R1)4a (ACTIVE STATE) == (R1)4b (INACTIVE)
WHEN ATP BINDS TO HEXAMERIZATION SITE
 CATALYTICALLY ACTIVE HEXAMERS (R1)6

89. Thioredoxin Physiologic reducing agent of RNR
Cys pair can swap H atoms with disulfide formed regenerate original enzyme
Thioredoxin gets oxidized to disulfide
Oxidized Thioredoxin gets reduced by thioredoxin reductase mediated
by NADPH (final electron acceptor)

90. Thymine Formation
Formed by methylating deoxyuridine monophosphate (dUMP)
UTP needed for RNA production, but dUTP not needed for DNA
If dUTP produced excessively, would cause substitution errors (dUTP for dTTP)
dUTP hydrolyzed by dUTP diphosphohydrolase to dUMP  methylated at C5 to form dTMP rephosphorylate to form dTTP

91. Chime Exercise
1DUD: dUTPase

92. Tetrahydrofolate (THF)
Methylation of dUMP catalyzed by thymidylate synthase
Cofactor: N5,N10-methylene THF
Oxidized to dihydrofolate
Only known rxn where net oxidation state of THF changes
THF Regeneration:
DHF + NADPH + H+  THF + NADP+ (enzyme: dihydrofolate reductase)
THF + Serine  N5,N10-methylene-THF + Glycine
(enzyme: serine hydroxymethyl transferase)

93. Anti-Folate Drugs
Cancer cells consume dTMP quickly for DNA replication
Interfere with thymidylate synthase rxn to decrease dTMP production
(fluorodeoxyuridylate – irreversible inhibitor) – also affects rapidly growing normal cells (hair follicles, bone marrow, immune system, intestinal mucosa)
Dihydrofolate reductase step can be stopped competitively (DHF analogs)
Anti-Folates: Aminopterin, methotrexate, trimethoprim

94. IN-CLASS QUESTION
IN von GIERKE’S DISEASE, OVERPRO- DUCTION OF URIC ACID OCCURS. THIS DISEASE IS CAUSED BY A DEFICIENCY OF GLUCOSE-6-PHOSPHATASE.
EXPLAIN THE BIOCHEMICAL EVENTS THAT LEAD TO INCREASED URIC ACID PRODUCTION?
WHY DOES HYPOGLYCEMIA OCCUR IN THIS DISEASE?
WHY IS THE LIVER ENLARGED?

95. ADENOSINE DEAMINASE DEFICIENCY
IN PURINE DEGRADATION, ADENOSINE  INOSINE
ENZYME IS ADA
ADA DEFICIENCY RESULTS IN SCID
“SEVERE COMBINED IMMUNODEFICIENCY”
SELECTIVELY KILLS LYMPHOCYTES
BOTH B- AND T-CELLS
MEDIATE MUCH OF IMMUNE RESPONSE
ALL KNOWN ADA MUTANTS STRUCTURALLY PERTURB ACTIVE SITE

96. ADA DEFICIENCY
IN-CLASS QUESTION: EXPLAIN THE BIOCHEMISTRY THAT RESULTS WHEN A PERSON HAS ADA DEFICIENCY
(HINT: LYMPHOID TISSUE IS VERY ACTIVE IN DEOXYADENOSINE PHOSPHORYLATION)

97. ADA DEFICIENCY ONE OF FIRST DISEASES TO BE TREATED WITH GENE THERAPY
ADA GENE INSERTED INTO LYMPHOCYTES; THEN LYMPHOCYTES RETURNED TO PATIENT
PEG-ADA TREATMENTS
ACTIVITY LASTS 1-2 WEEKS

98. Sugars
Pentoses (5-C sugars)
Numbering of sugars is “primed”

99. Sugars
D-Ribose and 2’-Deoxyribose
*Lacks a 2’-OH group

100. Nucleosides
Result from linking one of the sugars with a purine or pyrimidine base through an N-glycosidic linkage
Purines bond to the C1’ carbon of the sugar at their N9 atoms
Pyrimidines bond to the C1’ carbon of the sugar at their N1 atoms

101. Nucleosides

102. Phosphate Groups Mono-, di- or triphosphates
Phosphates can be bonded to either C3 or C5 atoms of the sugar

103. Nucleotides
Result from linking one or more phosphates with a nucleoside onto the 5’ end of the molecule through esterification

104. Nucleotides RNA (ribonucleic acid) is a polymer of ribonucleotides
DNA (deoxyribonucleic acid) is a polymer of deoxyribonucleotides
Both deoxy- and ribonucleotides contain Adenine, Guanine and Cytosine
Ribonucleotides contain Uracil
Deoxyribonucleotides contain Thymine

105. Nucleotides Monomers for nucleic acid polymers
Nucleoside Triphosphates are important energy carriers (ATP, GTP)
Important components of coenzymes
FAD, NAD+ and Coenzyme A

106. Naming Conventions Nucleosides: Nucleotides:
Purine nucleosides end in “-sine”
Adenosine, Cytosine
Pyrimidine nucleosides end in “-dine”
Thymidine, Guanidine
Nucleotides:
Start with the nucleoside name from above and add “mono-”, “di-”, or “triphosphate”
Adenosine Monophosphate, Guanidine Triphosphate, Deoxythymidine Diphosphate

107. In-Class Activities Look at the Nucleotide Structures
Take the Nucleotide Identification Quiz
Be prepared to identify some of these structures on an exam. Learn some “tricks” that help you to distinguish among the different structures