Nucleotide metabolism

Question 1

Ribose phosphate pyrophosphokinase

Answer 1

ribose-5-phosphate+ATP–>5’-phosphoribosyl-1-pyrophosphate (PRPP)+AMP

requires Mg²⁺

PRPP used for de novo synthesis and salvage of purines and pyrimidines, synthesis of NAD⁺, histidine biosynthesis, and conversion of guanine to GMP

Activated by P_i

Inibited by Purine ribonucleotides (A/G/I M/D/T P)

Question 2

Glutamine:phosphoribosyl pyrophosphate amidotransferase

Answer 2

PRPP (5’-phsophoribosyl-1-pyrophosphate) + Glutamine + H₂O—>5’-phosphoribosylamine + Glutamate + PP_i

Inhibitors: AMP, GMP, IMP

Activator: PRPP

First step of denovo purine synthesis; committed step; rate-limiting step

Question 3

De Novo Purine synthesis

Answer 3

Constructing a purine ring on a preformed ribose-5-phosphate (as PRPP)

Overall uses 4 ATPs

Uses Aspartate, CO₂, Glycine, Glutamine, N¹⁰-Formyl-tetrahydrofolate as sources of Carbons and Nitrogens

Tetrahydrofolate is especially important

End product is IMP

Question 4

PABA analogues

Answer 4

aka Sulfonaminde/Sulfa drug

structural analoge of para-aminobezonic acid, an intermediate in bacterial synthesis of folic acid–>competitvely inhibits synthetic pathway (Dihdropeteroate synthetase)

Folic acid (N10-Formyl-tetrahydrofolate) is a required cofactor in purine synthesis so de novo purine synthesis is slowed in these bacteria.

Humans cannot synthesize folic acid, so purine synthesis is not affected

Question 5

Folic Acid analogs

Answer 5

Including methotrexate and aminopterin (less common)

Inhibit reduction of dihydrofolate to tetraydrofolate catalyzed by dihydrofolate reductase

LImit amount of tetrahydorfolate (N10-Formyl-tetrahydrofolate) aroudn for purine synthesis thus slowing DNA replication

Used as an anti-cancer drug (toxic to all dividing cells), often used for leukemias

side effects: anemia, scaly skin, GI symptoms, respiratory tract disturbances, baldness

*Cancer cells can become resistant to MTX by amplification (duplication) of dihydrofolate reductase gene

Question 6

Trimethoprim

Answer 6

Inhibits tetrahydrofolate synthesis in prokaryotes

Inhibits bacterial dihydrofolate reductase

used as prophalatic and for UTIs

Question 7

Most common vitamin deficiency

Answer 7

Folic acid (B9)

-first seen as megaloblastic anemia–>Hb levels are low while blood marrow shows abnormally high number of megaloblastic cells (immature erythrocytes)

Question 8

AMP synthesis

Answer 8

used to synthesize AMP from IMP, requires GTP

Adenylosuccinate Synthetase:
IMP + Aspartate + GTP—>Adenylosuccinate + GDP + P_i

Inhbited by AMP

Adenylosuccinase: Adenylosuccinate—>AMP + fumarate

Question 9

Adenylosuccinate Synthetase:

Answer 9

Adenylosuccinate Synthetase:
IMP + Aspartate + GTP—>Adenylosuccinate + GDP + Pi

Inhbited by AMP

Question 10

Adenylosuccinase

Answer 10

Adenylosuccinate—>AMP + fumarate

Question 11

GMP synthesis

Answer 11

IMP dehydrogenase:
IMP + H2O + NAD+ ––>NADH + Xanthosine monophosphate (XMP)
inhibited by GMP and Mycophenolic Acid (deprives rapidly dviding B and T cells of nucleid acid components, used to prevent graft rejection)

GMP Synthetase:
XMP + ATP + Glutamine—>GMP + AMP + Glutamate

Question 12

IMP dehydrogenase:

Answer 12

IMP + H2O + NAD+ ––>NADH + Xanthosine monophosphate (XMP)

inhibited by GMP and Mycophenolic Acid (deprives rapidly dviding B and T cells of nucleid acid components, used to prevent graft rejection)

Question 13

Mycophenolic Acid

Answer 13

Mycophenolic Acid–>Inibits IMP dehydrogenase; deprives rapidly dviding B and T cells of nucleid acid components, used to prevent graft rejection

reversible, uncompetitive inhibitor

IMP + H2O + NAD+ ––>NADH + Xanthosine monophosphate (XMP)

also inhibited by GMP normally

Question 14

GMP Synthetase:

Answer 14

GMP Synthetase:
XMP + ATP + Glutamine—>GMP + AMP + Glutamate

Question 15

Purine Salvage:

Answer 15

2 enzymes:

Hypoxanthine-Guanine Phoshoribosyltransferase (HGPRT): Catalyzes 2 different rxns

Hypoxanthine + PRPP–>IMP + PP_i
Guanine + PRPP–>GMP + PP_i
(Lesch-Nyhan Syndrome)

Adenine Phosphoribosyltransferase (APRT)f

Question 16

Lesch-Nyhan Dyndrome

Answer 16

• X-linked recessive, near complete defciency of HGPRT
• inability to salvage hypoxanthine and guanine
• accumulation of uric acid (elevated in urine), end product of hypoxanthine and guanine degredation
• Orange crystals (uric acid) in diaper
• increased PRPP, decreased IMP and GMP; Increased de novo purine synthesis
• symptoms include severe mental retardation, self-mutilation, ivoluntary movements, gout

Question 17

Degredation of Purines

Answer 17

• Purines (and primidiens) are not essential components of the diet, most mononucleotides are synthesized de novo
• most purines degraded to uric acid rather than salvaged
• pyrimidies absorbed in intestinal mucosa dn returned to circulation (as are ribose and deoxyribose)

Question 18

Adenosice Deaminase (ADA)

Answer 18

Adenosine (nucleoSide) + H₂O––>Inosine + NH₃

Part of purine degredation (Inosine is a better substrate for purine nucleotide phosphorylase than adenosine)

Implicated in SCIDS

Question 19

SCIDs

Answer 19

Severe Combined Immunodeficiency

Deficiency of Adenosine Deaminase (ADA):
Adenosine–>Inosine

dATP accumulates in T-Cells and B-cells inhibiting DNA synthesis (inhibitor of Ribonucleotide reductase)

usually death before age 2 from infection

Question 20

Purine Nucleoside Phosphorylase

Answer 20

PNP or PNP(ase)

• Prefers Inosine or guanosine as substrates but can act on andenosine
• Purine nucleoside + P_i—>Purine + ribose-1-phosphate
• deficieny causes T-cell related immunodeficiencies; decreased uric acid production; increased purine nucleosides and nucleotides

Question 21

Xanthine Oxidase

Answer 21

2 reactions:

Hypoxanthine + O2 + H2O –––> Xanthine + H2O2

Xanthine + O2 + H2O –––> Uric Acid + H2O2

Can be inhibited by allopurinol

Question 22

Gout

Answer 22

• Hyperuricemia–caused most commonly by impaired elimination in urine but also by increased production due to errors in purine metabolism–leads to deposition of uric acid crystals in joints
• Uric acid crystals lead to acute arthritic joint inflammation
• Treated with Allopurinol: xanthine oxidase non-competitive inhibitor; accumulation and excretionof hypoxanthine and xanthine which are more soluble than uric acid
• Colchicine-microtubule inhibitor; produces clinical improvement but does not decres serum uric acid; probably inhibits migrations of white cells to joints
• 1° gout is genetic anf affects mainly males over 30
• 2° gout brought on by disorders including leukemia, polycythemia, HGPRT deficiency treatment of cancer with antimetabolites, chronic renal insufficiency; affects men and women of any age

Question 23

Pyrimidine synthesis

Answer 23

• pyrimidines synthesized before being attached to ribose-5-P (PRPP)
• Rate-limiting/committed step is CPS-II (Mammals: Inhibited by UTP, activated by ATP; Prokaryotes: aspartate transcarbamoylase is regulated step, inhibited by CTP)
• C’s and N’s come from Glutamine, Aspartic acid and CO₂ (no folic acid derivatives)
• Final product is UMP

Carbamoyl Phosphate synthetase II (CPS II)–>Aspartate transbarbamoylase–>dihydroorotase–>dihydroorotate dehydrogenase (uses NAD+)–>orotate phosphoribosyl-transferase–>OMP decarboxylase

Question 24

Carbamoyl Phosphate Synthetase II

Answer 24

2 ATP + CO₂ + Glutamine—->Carbamoyl Phosphate + 2 ADP + 2 P_i + Glutamate

Mammals: Inhibited by UTP, activated by ATP

Prokaryotes:

Question 25

CPS I vs CPS II

Answer 25

CPS I vs II •urea synthesis vs pyrimidine synthesis •free NH3 vs glutamine for N source •liver mitochondria vs cytosol of all nucleated cells •activated by N-acetylglutamate vs is not affected •no affect vs inhibited by UDP/UTP

Question 26

Orotic Aciduria

Answer 26

* Deficiency in orotate phosphoribosyltransferase and OMP decarboxylase (2 domains of a single polypeptide) * abnormal growth, megaloblastic anemia, excretion of large amounts of orotic acid in urine * (also caused by deficiency of OTC ins urea cycle (type II hyperammonemia) as excess carbamoyl phosphate gets shunted through pyramidine synthesis) * rare genetic form can be treated with dietary uridine despite utilization of dietary nucleotides, enough is absorbed * aquired form may appear in patients being treated for cancer with pyrimidine analog

Question 27

UMP--\>UTP

Answer 27

Uses NMP kinase and NDP kinase

Question 28

CTP Synthase

Answer 28

UTP + Glutamine + ATP----\>CTP + Glutamate + ADP + P_i

Question 29

Ribonucleotide Diphosphate Reductase

Answer 29

converts adenosine, guanosine, cytosine, and uridine ribonucleotides to deoxyribonucleotides; Inhibited by dATP and hydroxyurea NDP (except T) + Thioredoxin(2 S-H)---\>dNDP + thioredoxin(S-S) + H₂O Thioredoxin reductase: Thioredoxin(S-S; oxidized) + NADPH---\>Thioredoxin(2 S-H; oxidized) + NADP⁺ Enzyme contains a single active site 2 activitiy sites may bind ATP (activates) or dATP (inhibits) 2 substrate specificity sites bind NTPs and increase conversion of different species of ribonucleotieds to maintain proper balance (ATP, dATP, dTTP, dGTP)

Question 30

Thymidylate synthase

Answer 30

dUMP------\>dTMP N⁵, N¹⁰-methylTHF (B9) indirectly inhibited by MTX (methotrexate) Directly inhibited by 5-fluorouracil--\>(5-FdUMP) in cancer treatments

Question 31

Purine and Pyrimidine analogs used in treatment of cancer

Answer 31

* 5-fluorouracil: makes uracil analog; usually given with thymidine (protects normal, non-cancerous cells and cancer cells too kinda); converted to 5-fluorodeoxyridylic acid (dFUMP) which inhibits thymidilate synthase (affects cancer and non-cancer equally); in cancer cells 5-fluorocells is incorporated into RNA which is more detrimental to cancer cells than normal cells--\>the RNA is what really gets them * bromodeoxyuridine: thymidine analog (large group) * trifluorothyidine: thymidine analog * AraC

Question 32

Degradation fo Pyrmidines

Answer 32

* unlike purines, pyrimidines can be degraded to soluble precursors for other biomolecules * Can be salvaged by pyrimidine phosphoribosyltransferase (usese PRPP)