Lecture 1 part.1 Structure and Function of Nucleic Acid

Structure and function of nucleic acids
Purines & Pyrimidines; Nucleosides & Nucleotides
MOLECULAR BIOLOGY &
GENETICS
12.11, 2021

Building Blocks
 Nucleotides = Base + Sugar + Phosphate
 Nucleosides = Base + Sugar
 Nitrogen Bases
Purines (5 + 6 membered rings) – numbering
 AdenineGuanine
Pyrimidines (6 membered ring) – numbering
 Thymine Cytosine Uracil
 Pentose Sugars (numbering)
– Ribose
– Deoxy Ribose

5
 Planar, aromatic, and heterocyclic
 Derived from purine or pyrimidine
 Numbering of bases is “unprimed”
Nitrogenous Bases

DNA and RNA are composed of two different classes of
nitrogen containing bases: the purines and pyrimidines.
The most commonly occurring purines in DNA are adenine
and guanine:

8
A few of the modified nucleosides
that occur in tRNAs

Methylated Bases in E. coli
Typical sites of methylation include the N6 position of
adenine, the N4 position of cytosine, or the C5 position of
cytosine.
A characteristic feature of the sites of methylation, was that
they involved palindromic DNA sequences.

In addition to possessing a particular methylase, individual bacterial strains
also contained accompanying specific endonuclease activities. The
endonucleases cleaved at or near the methylation recognition site.
These specific nucleases, however, would not cleave at these specific
palindromic sequences if the DNA was methylated.
Restriction endonucleases
the 1978 Nobel Prize for Physiology or Medicine

Biological functions of nucleotides
1. Building blocks of nucleic acids (DNA and RNA).
2. Involved in energy storage, muscle contraction,
active transport, maintenance of ion gradients.
3. Activated intermediates in biosynthesis
(e.g. UDP-glucose, S-adenosylmethionine).
4. Components of coenzymes (NAD+, NADP+, FAD,
FMN, and CoA)
5. Metabolic regulators:
a. Second messengers (cAMP, cGMP)
b. Phosphate donors in signal transduction (ATP)
c. Regulation of some enzymes via adenylation and
uridylylation

Phospho-Anhydrides and Phosphate
Esters – High Energy Bonds

ATP- " biological currency"
 ATP is the universal energy currency of
the cell
 While ATP is one of four nucleotides
required for the synthesis of RNA, it is
primarily known in biochemistry for its role
in metabolism as the “biological currency"
of intracellular energy transfer.

Guanosine-5'-triphosphate (GTP)
GTP also has the role of a source of energy or an activator of
substrates in metabolic reactions, like that of ATP, but more
specific. It is used as a source of energy for protein
synthesis.
 GTP is essential to signal transduction, particularly with G-
proteins, in second-messenger mechanisms where it is
converted to GDP through the action of GTPases.
 GTP Hydrolysis Is Essential for Protein Import into the
Mitochondrial Matrix
 There are a very few instances in which GTP is used as a
phosphate donor or an energy source, most notable of which
is Tubulin, which must hydrolyze GTP to GDP to form the
microtubules of the cytoskeleton.

Glyceraldehyde-3-phosphate
Dehydrogenase
Phosphoglycerate Kinase
Enolase
PEP Carboxykinase
glyceraldehyde-3-phosphate
NAD+
+ Pi
NADH + H+
1,3-bisphosphoglycerate
ADP
ATP
3-phosphoglycerate
Phosphoglycerate Mutase
2-phosphoglycerate
H2O
phosphoenolpyruvate
CO2 + GDP
GTP
oxaloacetate
Pi + ADP
HCO3

+ ATP
pyruvate
Pyruvate Carboxylase
Gluconeogenesis
Summary of
Gluconeogenesis
Pathway:
Gluconeogenesis
enzyme names in
red.
Glycolysis
enzyme names in
blue.

Citrate cycle
CO
CH2
COO
COO
CH3CO~SCoA
C
CH2
COO
COO
CH2
HO
COO
C
CH
COO
COO
CH2 COO
CH
CH
COO
COO
CH2 COO
H2O
H2O
HO
CO2
CH2
CH2
COCOO
COO
CH2
CH2
COO
CO~ SCoA CO2
NAD+
NADH+H+
CH2
CH2
COO
COO
GDP+Pi
GTP
CH
CH2
COO
COO
OOC CH
C COO
H
HO
NAD+
NADH+H+
FAD
FADH2
H2O
acetyl CoA
H2O
oxaloacetate
citrate
synthase
citrate
aconitase
cis-aconitate
aconitase
isocitrate
NAD+
NADH+H+
isocitrate dehydrogenase
¦Á
-keto-
glutarate
¦Á
-ketoglutarate
dehydrogenase
complex
succinyl-CoA
ADP ATP
CoASH
succinyl CoA
syntetase
succinate dehydrogenase
fumarate
succinate
fumarase
malate
malate dehydrogenase
HSCoA
HSCoA

UTP
 UTP also has the role of a source of energy or an
activator of substrates in metabolic reactions, like that
of ATP, but more specific. When UTP activates a
substrate, UDP-substrate is usually formed and
inorganic phosphate is released. UDP-glucose enters
the synthesis of glycogen.
 Uridine is used for UDP-glucose, UDP-galactose, UDP-
mannose, etc., the building blocks of numerous
carbohydrates that are essential for many cellular
functions.

G
HK or GK
G-6-P
ATP ADP
G-1-P
UDPG
pyrophosphorylase
UDPG
UTP PPi Gn UDP
Gn+1
glycogen
synthase

CTP
 CTP is a high-energy molecule equal to ATP, but its role
in the organism is more specific than that of ATP. CTP is
used as the source of energy, and as a coenzyme in
metabolic reactions like the synthesis of
glycerophospholipids and glycosylation of proteins.
 CTP – very similar to UTP.
 However, instead of sugar, CTP is used with fats. CDP-
diacylglycerol, CDP-ethanolamine, and CDP-coline are
the building blocks of the phospholipids that make of the
cell membrane. Since all cell requires intact cell
membrane to survive, this is an exceedingly important
cellular function.

Adenine nucleotides are components of many
enzyme cofactors

NAD+ (Nicotinamide adenine dinucleotide) is a
coenzyme found in all living cells

in 1949, the American biochemists Morris Friedkin and Albert L.
Lehninger proved that NADH linked metabolic pathways such as the
citric acid cycle with the synthesis of ATP in oxidative phosphorylation
Nicotinamide adenine dinucleotide has several essential
roles in metabolism . It acts as a coenzyme in redox reactions,
as a donor of ADP-ribose moieties in ADP-ribosylation reactions,
as a precursor of the second messenger molecule cyclic ADP-
ribose, as well as acting as a substrate for bacterial DNA ligases
Function NAD+

NAD+/NADH, NADP+/NADPH
and FAD/FADH2
 Redox Cofactors

Function FAD
FAD is a prosthetic group in the enzyme
complex succinate dehydrogenase (complex
II) that oxidizes succinate to fumarate in the
eighth step of the citric acid cycle.
Another metabolic source of FADH2 is beta
oxidation, where FAD serves as a coenzyme
to acyl CoA dehydrogenase

Succinate dehydrogenase
 NAD used to oxidize oxygen-containing groups
 Aldehydes
 alcohols
 FAD used to oxidize C-C bonds

Summary of Glycolysis
ATP
ADP
Mg2+
PFK-1
GAP DHAP
glycerate
1,3-bisphosphate
NADH+H+
glyceraldehyde
3-phosphate
dehydrogenase
H3PO4
NADH+H+
NAD+
ADP
ATP
glycerate
3-phosphate
glycerate
2-phosphate
H2O
PEP
ATP
ADP
pyruvate kinase
lactate
pyruvate
G G-6-P F- 6-P F- 1,6-BP
NAD+
Phosphoglycerate
kinase
Isomerase
Aldolase
Mutase
Enolase
LDH
HK
ATP
ADP
Mg2+

Co-Enzyme A – Carrier for
Acetyl units in intermediary
metabolism, fatty acid
synthesis and oxidation

Function of Coenzyme A
 CoA (coenzyme A), notable for its role in the
synthesis and oxidization of fatty acids and the
oxidation of pyruvate in the citric acid cycle.
 Its main function is to carry acyl groups (such as
the acetyl group). A molecule of coenzyme A carrying an
acetyl group is also referred to as acetyl-CoA (where "A" stands for
acetylation). Acetyl CoA has a high acetyl group-transfer potential,
meaning that it carries an activated acetyl group, which it can deliver
for degradation and energy generation or for biosynthesis.

Regulatory/Signalling Molecules

cAMP
AMP can also exist as a cyclic structure known as cyclic AMP
(or cAMP). Within certain cells the enzyme adenylate cyclase
makes cAMP from ATP, and typically this reaction is regulated
by hormones such as adrenaline or glucagon. cAMP plays an
important role in intracellular signaling.
Rev Neurosci. 2005;16(1):23-41.
Function of cGMP-dependent protein kinases in the nervous system

Supplies ribose for
nucleotide metabolism
(to make nucleotides, NOT
to produce energy)
Where
does
the sugar
come
from?

Purine is
synthesized
from
amino acids,
tetrahydrofolate
and CO2
Sources of carbon and
nitrogen atoms in
the purine ring

Quantity and Quality of NA
 Bases are very nearly planar
 Aromaticity => large absorbance at 260nm
 Epsilon 260 ≈ 10,000 (M-1 cm-1 )
 The A260 ≈ 50 μg /ml for DS DNA
 The A260 ≈ 40 μg /ml for SS DNA or RNA
 Flat surfaces are hydrophobic
 Dipole-Dipole and Van Der Waals interactions
also stabilize stacked structures
 Bases have hydrogen bond donors and
acceptors
 H-bonding potential satisfied in paired structures

Purines are degraded in humans leads to uric acid
ADA GD

Uric Acid Excretion
 Humans – excreted into urine as insoluble
crystals
 Birds, terrestrial reptiles, some insects –
excrete insoluble crystals in paste form
 Excess amino N converted to uric acid
 (conserves water)
 Others – further modification :
Uric Acid  Allantoin  Allantoic Acid  Urea  Ammonia

Trinucleotide repeat disorders
SNP
altered AMP deaminase (AMPD) activity strategy
-evaluated in experimental models of heart
failure, ischemic heart disease,
ischemia/reperfusion injury and type 2 diabetes.

Deleterious consequences of defective
purine metabolism
• Gout (excess accumulation of uric acid)
• Loss of regulation of purine nucleotide synthesis
• The fact that, in man, purine degradation is already
terminated with uric acid can lead to the problems. Man
metabolizes 400 – 600 mg/per day.
• Xanthine oxidase is the point for pharmacological check
in patients with hyperuricemia (gout)
• Hyperuricemia – level of urates in blood is higher than
its Product of solubility, therefore urates crystallize in
soft tissues and joints forming deposits called tophi,
causing an inflammatory reaction - acute gouty arthritis
→ chronic gouty arthritis

What is SNP ?
 A SNP is defined as a single base change
in a DNA sequence that occurs in a
significant proportion (more than 1 percent)
of a large population.
• SNPs are the most simple form and most
common source of genetic polymorphism in the
human genome (90% of all human DNA
polymorphisms).

Some Facts
 In human beings, 99.9 percent bases are
same.
 Remaining 0.1 percent makes a person
unique.

2007 Scientific Breakthrough
of the Year

Single (Simple) Nucleotide
Polymorphisms (SNPs)
 SNPs are used for identification and forensics
 SNPs are used for mapping and genome-wide association
studies of complex diseases
 SNPs are used for estimating predisposition to disease
 SNPs are used for immigration & citizenship in the UK
 SNPs are used to predict specific genetic traits
 SNPs are used for classifying patients in clinical trials
GCTGTATGACTAGAAGATCGAT
GCTGTATGACGAGAAGATCGAT

Lecture 1 part.1 Structure and Function of Nucleic Acid

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Lecture 1 part.1 Structure and Function of Nucleic Acid