The 5’ phosphate end of the new nucleotide is linked to the 3’-OH of the last nucleotide

of the growing chain by DNA polymerase III.

Aspect Description

DNA and RNA strands have directionality with a 5’ (five-

Directionality of prime) end and a 3’ (three-prime) end. The 5’ end has a
DNA and RNA phosphate group attached to the fifth carbon of the sugar.
Strands The 3’ end has a hydroxyl group attached to the third
carbon.

Transcription is the process of copying a segment of DNA

into RNA by RNA polymerase. RNA polymerase reads the
DNA template strand in the 3’ to 5’ direction and
Transcription
synthesizes RNA in the 5’ to 3’ direction. RNA polymerase
can only add nucleotides to the 3’ end of the growing RNA
strand.

Translation is the process of using the mRNA sequence to

direct protein synthesis. The ribosome reads mRNA in the
Translation
5’ to 3’ direction. The directionality ensures that codons
are read in the correct order for proper protein assembly.

Enzymes involved in transcription and translation ensure

Enzyme unidirectional processes. RNA polymerase synthesizes
Specificity RNA in the 5’ to 3’ direction. Ribosomes move along
mRNA in the 5’ to 3’ direction during translation.

- Ensures accurate expression of genetic information. <br>

Importance of
- Maintains proper function of cells and organisms by
Directionality
ensuring correct protein synthesis.
DNA replication:
A promoter is a DNA segment that initiates gene transcription. A promoter is a
sequence of DNA needed to turn a gene on or off. The process of transcription is
initiated at the promoter. Usually found near the beginning of a gene, the promoter has a
binding site for the enzyme used to make a messenger RNA (mRNA) molecule. It is
typically between 100 and 1,000 bases long and is located next to genes on
chromosomes. The base sequence of a promoter allows RNA polymerase to bind,
along with various proteins known as transcription factors.

• Repressor sequences within the promoter bind transcription factors that

prevent RNA polymerase from binding, thus stopping gene transcription.

• Activator sequences within the promoter bind transcription factors that

encourage RNA polymerase binding, leading to gene transcription.

Cells can change their pattern of gene expression depending on what transcription
factors are produced to bind with the promotors. Some groups of genes have very
similar promoters, so they are all expressed together even if they are located on
different chromosomes. Some transcription factors only bind to the promoter after
combining with another molecule. For example, the receptor for testosterone acts as a
transcription factor. If testosterone binds to it, the receptor can bind to many promoters
of genes with roles in male sexual development and activity.

Concept Description

Regulation by Two groups of proteins mediate binding of RNA

Proteins polymerase to the promoter.

Transcription
Form a complex with RNA polymerase at the promoter.
Factors

RNA Polymerase Cannot initiate transcription without transcription

Dependency factors; their levels regulate gene expression.

Bind to DNA sequences outside of the promoter and

Regulatory Proteins
interact with transcription factors.

Bind to enhancer sites and increase the rate of

Activator Proteins
transcription by mediating complex formation.

Bind to silencer sequences and decrease the rate of

Repressor Proteins
transcription by preventing complex formation.

Tissue-Specific Presence of certain transcription factors or regulatory

Factors proteins may be tissue-specific.

Hormones and other chemical signals can moderate

Chemical Signals
protein levels and mediate changes in gene expression.
The DNA sequences that regulatory proteins bind to are called control elements

▪ Some control elements are located close to the promoter (proximal elements)
while others are more distant (distal elements)

▪ Regulatory proteins typically bind to distal control elements, whereas

transcription factors usually bind to proximal elements

▪ Most genes have multiple control elements and hence gene expression is a
tightly controlled and coordinated process

• Only some DNA sequences code for polypeptides. In humans, only about 1%-2%
of DNA codes for a polypeptide. These coding regions are called exons. The
remainder of the DNA is non-coding. The non-coding regions are typically made
up of repetitive sequences of DNA:

Production of RNA Some regions on DNA function to produce tRNA and rRNA

Gene expression Non-coding regions can have an role in regulating the expression of
genes by promoting or inhibiting.

Telomeres Telomeres are located on the ends of eukaryote chromosomes, they

have a protective function because DNA cannot be replicated all the
way to the ends, so telomeres prevent loss of important genes. Also
protects the DNA from degradation during replication
Summary of common types of regulating proteins and associated sequences found in
eukaryotes.

DNA Binding
Function
Sequence protein

Enhancers Activator Activator proteins bind to enhancer sequences of

DNA to greatly increase the rate of transcription
of a gene.

Operator Repressor Repressor proteins bind to non-coding regions of

(Silencers) DNA to either block or reduce the transcription of
a gene.

Promoter RNA A region of DNA located close to a specific gene.

Polymerase Once bound to the sequence RNA polymerase
transcribes the gene.
The separation of transcription and translation into separate compartments in
eukaryotic cells allows for significant post-transcriptional modification to occur before
the mature mRNA exits the nucleus. RNA is modified by adding extra nucleotides to give
each end of mRNA a special structure.

• Five-prime caps—a modified nucleotide is added to the 5′ end of the RNA. This
nucleotide has three phosphate groups instead of one, so it is similar to ATP. Its base is
guanine, with an extra methyl group added.

• Poly (A) tails—between 100 and 200 adenine nucleotides are added to the 3′ end of
the RNA. Translation stops before the ribosome reaches the poly (A) tail. The 5′ cap and
the poly (A) tail both stabilize the ends of the mRNA by protecting them from digestion
by nuclease enzymes. They also encourage further modification of the original
transcript.

Eukaryotic genes that code for polypeptides contain two types of sequence:

• exons—coding sequences that are expressed by translation into the amino acid
sequence

• introns—intervening sequences that are not expressed.

Introns are mostly between 20 and 200 nucleotides long. The mean number of introns
per gene in humans is 7.8 and the mean number of exons is 8.8.

RNA transcribed from genes coding for polypeptides contains alternating exons and
introns.

The introns are removed and digested into single nucleotides. The remaining exons are
spliced together to produce an uninterrupted base sequence for translation. The mRNA
is then mature and can be exported from the nucleus for translation by ribosomes in the
cytoplasm.

Alternative splicing increases the diversity of the proteome—all of the proteins

expressed in an organism.

Alternative splicing is a process that allows a single gene to produce multiple variants
of a polypeptide. This is achieved by modifying the primary RNA transcript in different
ways after transcription. The most common method of alternative splicing is exon
skipping, where certain exons (coding regions) are included or excluded from the final
mRNA. This results in different mRNA variants, which then translate into different
polypeptides.
 • Primary Transcript: The initial RNA copy made from the DNA template is the
same for all variants.

• Post-Transcriptional Modification: Differences arise during the processing of

this RNA, particularly in how exons are spliced together.

• Exon Skipping: The most frequent change, where some exons are skipped in
some mRNA variants but included in others.

• Functional Diversity: This process allows for the production of polypeptides

with different forms and functions from a single gene, increasing the protein
diversity without needing additional genes.

The immunoglobulin genes undergo alternative splicing to produce different forms of

antibodies, such as membrane-bound and secreted forms. This splicing allows for the
generation of diverse antibody molecules from a single gene, which is essential for the
immune system’s ability to recognise a wide array of antigens.

Another common example of alternative splicing in humans is the fibronectin gene.

Fibronectin is a high-molecular-weight glycoprotein that plays a crucial role in cell
adhesion, growth, migration, and differentiation. The fibronectin gene undergoes
alternative splicing to produce different isoforms that are either soluble or insoluble,
depending on the tissue type and developmental stage.

The soluble form of fibronectin is found in the blood plasma, while the insoluble form is
part of the extracellular matrix. This versatility allows fibronectin to perform various
functions in different cellular environments, contributing to processes like wound
healing and embryonic development.

The Drosophila DSCAM gene. This gene is involved in the development of the nervous
system in fruit flies and can produce over 38,000 different protein isoforms through
alternative splicing. This diversity is crucial for the proper wiring of the nervous system,
ensuring that neurons connect correctly.
Another example of alternative splicing is the troponin T gene in heart muscle, which
produces different variants of the troponin T protein, crucial for muscle contraction.

Exons 4 and 5 are alternatively spliced to produce four different versions of troponin C:

• TnT1: all exons present in the mature mRNA

• TnT2: exon 5 present but not exon 4

• TnT3: exon 4 present but not exon 5

• TnT4: neither exon 4 nor exon 5 present.

Exons 4 and 5 both make troponin C more sensitive to calcium ions.

• mRNA translated in a 5' to 3' direction
• binding of ribosome to mRNA; small sub-unit then large subunit.Ribosome has 2
t-RNA binding sites (P-site and A-site)
• tRNA-activating enzymes link correct amino acid to each tRNA; (activated) tRNA
has an anticodon and the corresponding amino acid attached at 3’ CCA;
• first tRNA (within anticodon UAC) carrying Met binds to start codon AUG on
mRNA in P-site
• second tRNA with amino acid binds to ribosome in A-site.
• Enzyme peptidyl transferase is involved in peptide bond formation between the 2
amino acids and requires energy
• large subunit moves down mRNA BY 1 codon 5’ to 3’
• amino acid (or polypeptide) on first tRNA is transferred after it forms peptide
bond with amino acid on second tRNA (translocation)
• small subunit of ribosome moves down the mRNA to follow large subunit
• loss of tRNA (from E-site) from 1st site and new tRNA binds in A-site
• Finally ribosome reaches a stop codon / termination (UAA, UAG, UGA).
When a stop codon is reached translation is stopped:
 • a release factor attaches to the A site

• the polypeptide chain is released

• the ribosome complex dissembles ready for reuse translating another

mRNA molecule

Modification Type Description

Removal of Removal of the amino acid methionine from the 5′ end of the
Methionine polypeptide, where it was placed during initiation of translation.

Changes to the side chains of amino acids in the polypeptide, such

Changes to Side
as phosphorylation or addition of carbohydrate molecules, can
Chains
produce over 100 chemically different amino acids in proteins.

Folding the polypeptide and making intramolecular interactions to

Folding and
stabilize the tertiary structure, e.g., disulfide bonds formed between
Stabilization
cysteines that come close together due to folding.

Conversion of Conversion of propeptides into mature peptides by removal of part of

Propeptides the polypeptide chain.

Formation of
Combining two or more polypeptides into the quaternary structure
Quaternary
proteins.
Structure

Addition of Non-
Combining non-polypeptide components into the quaternary
Polypeptide
structure of conjugated proteins.
Components
Modification of preproinsulin to insulin:

1. Transcription: The insulin gene (INS) on chromosome 11 is transcribed in

pancreatic beta cells to produce mRNA.

2. Translation: The mRNA is translated by ribosomes on the rough endoplasmic

reticulum (RER), producing a 110-amino acid polypeptide called preproinsulin.

3. Signal Peptide Removal: Preproinsulin enters the RER lumen, where a protease
removes a 24-amino acid signal peptide from the N-terminal, resulting in an 86-
amino acid chain called proinsulin.

4. Folding: Proinsulin folds into its tertiary structure, stabilized by three disulfide
bonds.

5. Cleavage: Proteases break peptide bonds at two positions in proinsulin, excising

a 33-amino acid sequence and leaving two chains: the A-chain (21 amino acids)
and the B-chain (initially 32 amino acids), held together by disulfide bonds.

6. Final Modification: A protease removes a pair of amino acids from the C-

terminal of the B-chain, yielding mature insulin with a total of 51 amino acids.
Proteasomes play a crucial role in removing non-functional and damaged proteins from
the cell while recycling amino acids for new protein synthesis.

Recycling of Amino Acids by Proteasomes

Most proteins produced by translation have a relatively short lifespan in the cell for
several reasons:
 • Changing Cell Activities: Proteins may no longer be needed as the cell
progresses through different stages of the cell cycle.

• Structural Changes: Proteins can be altered by free radicals or other reactive

chemicals, rendering them non-functional.

• Misfolding or Denaturation: Proteins can become misfolded or denatured in

various ways.

Proteasome Structure and Function:

• Regulatory Subunits: Located at both ends of the proteasome, these subunits

recognize proteins tagged with ubiquitin and queue them for digestion.

• ATP-Fueled Subunit: This subunit unfolds proteins and feeds them into the
proteasome core for digestion.

• Proteasome Core: A hollow cylindrical structure containing proteases with

active sites facing the inner chamber, where tagged polypeptides are digested
into short chains of amino acids.

Process of Protein Degradation:

1. Ubiquitination: Proteins marked for degradation are tagged with ubiquitin,

signaling proteasomes to digest them.

2. Recognition and Unfolding: Ubiquitinated proteins are recognized by regulatory

subunits and unfolded by an ATP-fueled subunit.

3. Digestion: The unfolded proteins are fed into the proteasome core, where
proteases break them down into short amino acid chains.

4. Amino Acid Recycling: These short chains are further digested in the cytoplasm
to yield amino acids, which can be reused for new protein synthesis.