NUCLEIC ACID

Lecture | Week 2

NUCLEIC ACIDS: DNA STRUCTURE - Guanine can bond specifically only with
 Double Helix – 2 strands Cytosine, Adenine can bond only with
 Polymer - consists of multiple monomers Thymine
o Polynucleotide = Nucleic Acid - We called these pairings complementary
base pairs.
 A guanine-cytosine pair is held by three
hydrogen bonds
 An adenine-thymine pair is held by two
hydrogen bonds.
 The two DNA strands are antiparallel, that is,
their 5' - 3' orientation runs in opposite directions
(Chargaff).

 If a molecule is composed of a purine or

Nucleotide Structure pyrimidine base and a ribose or deoxyribose
sugar, this chemical unit is called a
 Pentose Sugar - Deoxyribose nucleoside.
 Nitrogenous Base Pair:  When the phosphate group of one nucleotide
- Adenine bonds with the sugar group of another
- Guanine nucleotide, they form a phosphodiester bond.
- Thymine This process results in the formation of a long
- Cytosine chain of bonded nucleotides. Such a chain is
referred to as a polynucleotide, and
polynucleotides make up nucleic acids, such
as DNA and RNA.

NUCLEIC ACIDS: RNA STRUCTURE

 Single Strand Nucleic Acid
 Pentose Sugar: Ribose
 Nitrogenous Base Pair:
- Adenine
 Sugar - Phospate bond between nucleotides will
- Guanine
form a one long chain, that will serve as the
- Uracil
backbone of the DNA strand. It is also called a
- Cytosine
phosphodiester bond.
 Several kinds of RNA play an important role in
 Purines (adenine and guanine) are two-carbon
cellular activities. Three major classes of cellular
nitrogen ring bases.
RNA molecules function during the expression
 Pyrimidines (cytosine and thymine) are one-
of genetic information:
carbon nitrogen ring bases.
- ribosomal RNA (rRNA),
 When nitrogenous bases pair together, it is
- transfer RNA (tRNA),
called base pairing.
- messenger RNA (mRNA)
 When 2 nitrogenous bases pair together, they
 Less stable due to the presence of the 2'
form hydrogen bonds that hold the two strands
hydroxyl group, making it more susceptible to
together.
hydrolysis.
 In general, for DNA (Chargaff):

NUCLEIC ACIDS: RNA FORMS

1. Ribosomal RNA (rRNA)

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 - Ribosomal RNA usually constitutes about process called DNA denaturation, or DNA
80% of all RNA in E. coli cells. melting.
- They are important structural components of  On the other hand, when the temperature of a
ribosomes, which function as non-specific DNA solution is low enough, the hydrogen
sites of protein synthesis during translation. bonds form again, and finally, all hydrogen
2. Transfer RNA (tRNA) bonds are restored: DNA renaturation.
- accounts for up to 15% of the RNA in a
typical cell. Primary & Secondary Structure of DNA
- It carries amino acids to the ribosome during
translation, aiding in the translation of DNA
to mRNA to protein. PRIMARY SECONDARY
3. Messenger RNA (mRNA).
Phosphodiester Bond Double Helix & Anti-
- carry genetic information from the DNA of
Backbone parallel strand
the gene.
- The mRNAs vary in size, reflecting the
range in the sizes of the proteins encoded
by the mRNA.

NUCLEIC ACIDS: DNA vs RNA

DNA RNA

SUGAR Deoxyribose Ribose

BASE PAIR G=C G=C

A=T U=T Tertiary Structure of DNA
 The tertiary structure of DNA involves
# OF STRAND Double strand Single strand
supercoiling, where the already twisted double
LOCATION Nucleus Cytoplasm helix is further twisted to fit inside the cell, much
like twisting an already twisted ladder even
STABILITY More stable Less stable more.
 There are two types of supercoiling:
- positive supercoiling, which twists in the
PHYSICAL PROPERTIES OF NUCLEIC ACIDS
same direction as the double helix,
- negative supercoiling, which twists in the
Denaturation of DNA opposite direction.
 An important feature of double helix DNA is the  In eukaryotes, DNA is organized into chromatin
ability to separate the two strands (a process structure for further compaction.
called denaturation)  DNA wraps around proteins called histones to
 To base pair the two strands together, a process form nucleosomes, creating a "beads on a
called renaturation, without disrupting the string" appearance.
covalent bonds that make up the sugar-  These nucleosomes then coil further to form a
phosphate backbone. more compact structure known as chromatin.
 The hydrogen bonds can be disrupted by:
- high temperature Secondary Structure of RNA
- low salt concentration
1. Stability and Protection:
- high pH in vitro
- Secondary structures like hairpins, bulges,
 For example, when a DNA solution is heated
and internal loops help stabilize RNA
enough, the hydrogen bonds that hold the two
molecules and protect them from
strands together weaken and finally break, a
degradation by nucleases.
2. Regulation of Gene Expression:

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 - Hairpin structures can regulate gene  Enzymes Involved: DNA polymerases are key
expression by affecting the accessibility of enzymes that synthesize new DNA strands by
RNA to ribosomes and other proteins adding nucleotides to the existing DNA template.
involved in translation.
3. Functionality of Non-Coding RNAs: Transcription
- Non-coding RNAs, such as tRNA and rRNA,  The process of copying a segment of DNA into
rely on specific secondary structures for RNA.
their function. For example, tRNA’s  Location: Takes place in the nucleus of
cloverleaf structure is essential for its role in eukaryotic cells and in the cytoplasm of
translation. prokaryotic cells.
 Key Enzyme: RNA polymerase binds to the
Tertiary Structure of RNA DNA at a specific region called the promoter and
1. Catalytic Activity: synthesizes RNA by adding complementary
- Ribozymes, RNA molecules with catalytic RNA nucleotides to the growing RNA strand.
activity, depend on their tertiary structures to  Product: The primary product is messenger
form active sites for biochemical reactions. RNA (mRNA), which carries the genetic
The folding of RNA into specific three- information from DNA to the ribosome, where
dimensional shapes is crucial for their proteins are synthesized. Other types of RNA
enzymatic functions. (tRNA, rRNA) are also transcribed from DNA.
2. Structural Roles:
- Tertiary structures in rRNA are essential for Translation
the formation and function of the ribosome,  The process by which the genetic code carried
the cellular machinery for protein synthesis. by mRNA is decoded to produce a specific
3. RNA-Protein Interactions: protein.
- Tertiary structures enable RNA to interact  Location: Occurs in the cytoplasm at the
specifically with proteins, forming ribosome.
ribonucleoprotein complexes necessary for  Key Players:
processes such as splicing (e.g., - mRNA: Carries the genetic information from
spliceosomes) and translation (e.g., DNA to the ribosome.
ribosomes). - tRNA: Brings the appropriate amino acids to
4. Regulatory Functions: the ribosome. Each tRNA has an anticodon
- Tertiary structures of regulatory RNAs (e.g., that matches the codon on the mRNA.
miRNAs and siRNAs) facilitate their - Ribosome: The molecular machine that
interactions with target mRNAs, guiding facilitates the assembly of amino acids into a
gene silencing and regulation. polypeptide chain based on the sequence of
codons in the mRNA.
CENTRAL DOGMA  Product: A polypeptide chain that will fold into a
 Is the process in which genetic information flows functional protein.
from DNA to RNA, to make a functional product
protein.

Replication
 The process by which DNA makes an identical DNA
copy of itself. REPLICATION
 Location: Occurs in the nucleus of eukaryotic  Follows the Semiconservative Replication
cells. Mechanism
- Each of the two strands of the DNA double
helix serves as a template for the production
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 of a new, complementary strand. As a result, of the replication fork. This discontinuous
each newly formed DNA molecule consists synthesis occurs because DNA
of one original (parental) strand and one polymerase can only add nucleotides to
newly synthesized strand. the 3' end.

Termination
THREE DIFFERENT PHASES OF
1. RNAse H - It recognizes and removes the RNA
REPLICATION
primers
2. After RNase H removes the RNA primer, DNA
Initiation polymerase fills in the resulting gaps with DNA
1. DNA replication starts at specialized sites called nucleotides, using the adjacent Okazaki
origins of replication. fragment as a primer.
2. Helicase 3. Once the RNA primers are removed and
- Unwind the DNA double helix, creating two replaced with DNA, DNA ligase seals the nicks
single strands of DNA. between the newly synthesized DNA fragments,
- The unzipping enzyme in DNA replication. creating a continuous DNA strand.
3. This unwinding creates a replication fork, a Y-
shaped region where the DNA is split into two DNA MUTATION, DAMAGE, MISMATCH &
separate strands.
REPAIR

DNA MUTATION
 a DNA molecule that occurs when changes
escape the cell's proofreading and repair
mechanisms. These genetic changes can affect
a single nucleotide pair or larger segments of a
chromosome and can have various effects on an
organism's traits and functions.
4. Single-Strand DNA Binding Proteins (SSBs)
- When the two strands are separated, single-
Types of Mutations
strand DNA binding proteins (SSBs) bind
1. Point Mutation (Substitution)
selectively to single-stranded DNA as soon - Missense mutation
as it forms. The binding of SSBs can - Nonsense mutation
stabilize the single-stranded DNA so they - Silent mutation
will not anneal to reform the double helix 2. Insertion and Deletion
3. Tautomeric Shifts
Elongation
1. DNA Primase - is responsible for copying a Point Mutation
short stretch of the DNA template strand to  Arises when a base pairs with an inappropriate
produce the RNA primer sequence or an RNA partner during DNA replication.
complementary strand.  Types of Point Mutations
2. DNA Polymerase - extends the RNA primer by - Silent Mutation: The altered codon still
adding nucleotides complementary to the codes for the same amino acid, so the
template strand. This occurs in a 5' to 3' protein remains unchanged.
direction. - Missense Mutation: The altered codon
* Synthesis is continuous on the leading codes for a different amino acid, potentially
strand. changing the protein's function.
* Synthesis is discontinuous on the lagging - Nonsense Mutation: The altered codon
strand becomes a stop codon, leading to a
truncated and usually nonfunctional protein.
Okazaki fragments
 The strand that is synthesized
discontinuously, opposite to the direction
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 - Amino form of adenine (A) and cytosine (C)
- Keto form of guanine (G) and thymine (T)

DNA MISMATCH REPAIR

Excision of the Error-containing DNA

 The newly synthesized DNA strand containing
the error is identified and nicked. MutH nicks the
strand at a hemimethylated GATC site.
Insertion and Deletion  An exonuclease removes a segment of the DNA
 Insertion: An additional base pair is inserted strand, including the mismatch.
into the DNA sequence.
 Deletion: A base pair is removed from the DNA Synthesis of New DNA
sequence.
 DNA polymerase fills in the gap created by the
 These are called frameshift mutations
exonuclease, using the undamaged, original
because the frame of the triplet reading during
DNA strand as a template.
translation is altered.

Ligation
 DNA ligase seals the newly synthesized
segment into the existing strand, restoring the
integrity of the DNA molecule.

Recognition of Mismatch
 Specialized proteins recognize and bind to
mismatched bases. In prokaryotes, the MutS
protein detects the mismatch. In eukaryotes, the
homologous MutSα complex (composed of
MSH2 and MSH6 proteins) carries out this
function.

Recruitment of Repair Proteins

 Once the mismatch is recognized, additional
proteins are recruited to the site. In prokaryotes,
MutL (which interacts with MutS) and MutH
(which nicks the newly synthesized strand) are
recruited. In eukaryotes, MutLα (composed of
MLH1 and PMS2 proteins) is recruited to the
mismatch site.

Tautomeric Shift DNA DAMAGE

 Tautomeric shifts are a specific type of mutation  In addition to experiencing those spontaneous
that occur when a nucleotide base in DNA mutations caused by misreading the genetic
temporarily changes its chemical form, or code, organisms are frequently exposed to
tautomer, leading to incorrect base pairing mutagens that cause damage to DNA.
during DNA replication. This can result in point  DNA damage is a chemical alteration to DNA
mutations if the altered base pairs with the which can be introduced by many different ways.
wrong partner. If the DNA damage is left unrepaired, it can lead
 Tautomeric Forms: Nucleotide bases can exist to mutation.
in alternative chemical forms called tautomers.  Some common examples of DNA damage are:
The common tautomers in DNA are:

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 - base modifications caused by base analogs
and alkylating agents,
- pyrimidine dimers caused by ultraviolet (UV)
radiation, DNA Damage: UV Radiation
- free radicals caused by ionizing radiation.  Mechanism: UV radiation causes covalent
bonding between adjacent pyrimidine bases
DNA Damage: Base Analogs (thymine or cytosine) on the same DNA strand.
Mechanism  The shape of the DNA of both produces is
 What They Are: Base analogs are chemicals distorted, and thus, both replication and
that look a lot like the normal building blocks of transcription are interfered with.
DNA (A, T, C, G).
 What They Do: Because they resemble these
building blocks, they can sneak into the DNA
during replication (when DNA is being copied).
 The Problem: Once in the DNA, base analogs
can sometimes pair up incorrectly with other
bases, leading to mistakes in the DNA
sequence.

DNA Damage: Ionizing Radiation

 Production of Reactive Oxygen Species (ROS):
When ionizing radiation hits water molecules in
cells, it splits the water molecules into highly
reactive oxygen species (ROS) such as hydroxyl
DNA Damage: Alkylating Agents radicals.
 DNA Damage: These ROS are very reactive and
Mechanism
can cause various types of damage to DNA,
 What They Are: Alkylating agents are
including:
chemicals that add small groups called alkyl
o Base Modifications: Changing the
groups (like methyl or ethyl) to the DNA bases.
 What They Do: These added groups change chemical structure of DNA bases.
the shape and structure of the DNA bases, o Single-Strand Breaks (SSBs): Breaking
making them pair incorrectly with other bases one of the two strands of the DNA
during replication. double helix.
o Double-Strand Breaks (DSBs): Breaking
both strands of the DNA double helix.

DAMAGE REPAIR: SINGLE-STRAND DAMAGE

REPAIR
 Single-strand DNA damage repair is a process
that specifically addresses and fixes damage
affecting only one strand of the DNA double
helix.
1. Direct Repair
2. Excision Repair System
o Base Excision Repair
o Nucleotide Excision Repair

Direct Repair

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Mechanism:
 Direct Reversal: Some types of DNA damage
can be directly reversed by specific enzymes
without the need for excision or replacement of
nucleotides.
 Example:
o Photoreactivation: In some organisms,
an enzyme called photolyase uses
energy from visible light to break the
bonds of pyrimidine dimers (caused by
UV radiation), directly reversing the
damage.

Excision Repair System

 Base Excision Repair
- This type of damage often involves
modifications to individual DNA bases
 Nucleotide Excision Repair
- is a DNA repair mechanism that fixes bulky,
helix-distorting lesions caused by
environmental factors such as UV radiation
and chemical exposure.

DAMAGE REPAIR: DOUBLE-STRAND DAMAGE

REPAIR
 Double-Strand Break (DSB) Repair is a critical
cellular process that fixes breaks in both strands
of the DNA double helix.
 Such breaks are particularly dangerous because
they can lead to significant genomic instability
and are associated with various diseases,
including cancer.
 There are two primary mechanisms for repairing
DSBs:
1. Homologous Recombination (HR)
2. Non-Homologous End Joining (NHEJ).

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