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Name: Jason Lloyd Fantonalgo Score:

Nixon Rauff Fowler Date: November 30, 2023
Manuel Castones Legarde III

EXERCISE NO. 1

PROKARYOTIC AND EUKARYOTIC REPLICATION, TRANSCRIPTION AND

TRANSLATION

INTRODUCTION

OBJECTIVES

The students should be able to:

1. construct models for prokaryotic replication, transcription and translation

using plastic chips or other easily accessed materials;
2. relate the colored plastic chips to different molecular components that figure
out in prokaryotic and eukaryotic replication, transcription and translation;
3. elucidate prokaryotic and eukaryotic replication, transcription and translation
using simple materials.

MATERIALS

(depends on what materials you will use)

PROCEDURE

A. CONSTRUCTION OF Models for DNA Replication

1. Using your collected materials, design a model each for prokaryotic and eukaryotic
replication.
2. Include all elements that converge at the replication fork during DNA replication for
both prokaryotes and eukaryotes.
3. Group the plastic chips according to color. One color represents one specific
replisome component.
4. Design your replication fork models and place the colored chips in their
corresponding places.
5. Make a legend of your colored chips for reference purposes.
6. Take pictures of your models and include these in your final reports.

B. CONSTRUCTION OF Models for DNA Transcription and Translation

1. Using the same set of materials, construct models for prokaryotic and eukaryotic
initiation of transcription/translation and post-transcriptional processing (in
eukaryotes).
2. Do the same for this exercise.
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SKETCHES

LEGEND:
Violet = Replicated strand
White = Topoisomerase
Blue = Clamp
Green = Polymerase
Red = Helicase
Pink = Primer
Yellow = Parent Strand

Black = SSBP

Light Blue = A-T

Light Green = G-C

Prokaryotic Replication Fork

LEGEND:
Violet = Replicated strand
White = Topoisomerase
Blue = Clamp
Green = Polymerase
Red = Helicase
Pink = Ligase
Yellow = Parent Strand

Orange = Primer

Black = SSBP

Light Blue = A-T

Light Green = G-C

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QUESTIONS

1. Discuss main points of difference between

a. Prokaryotic and Eukaryotic replication
- Prokaryotes have one origin of replication bidirectionally. On the other
hand, Eukaryotic replication has bidirectional multiple origins of
replication.
- Eukaryotic replication relies on a comparatively straightforward system
with a few essential enzymes like DNA helicase, DNA polymerase, and
DNA ligase. In contrast, prokaryotic replication employs a more intricate
and multifaceted machinery, encompassing numerous enzymes and
proteins, which collectively contribute to the efficiency and complexity of
the replication process in these microorganisms.

b. Leading and Lagging Strand Synthesis

- Leading strand synthesis is continuous and occurs in the same direction as
the replication fork moves while lagging strand synthesis is discontinuous
and occurs in the opposite direction of the replication fork moves. The two
strands are ultimately joined together to form a continuous DNA molecule.
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c. Replication fork components of Prokaryotes and Eukaryotes

- Prokaryotes are circular DNA, bidirectional, and only have one point of
origin, while Eukaryotes are linear DNA, unidirectional, and have multiple
points of origin.

d. Prokaryotic and Eukaryotic Transcription

- Prokaryotes possess nucleoid instead of nucleus, have only one RNA
polymerase, have simpler promoters, and often lack introns. On the other
hand, eukaryotes possess distinct nuclei, have multiple RNA polymerases,
have complex promoters, and have introns that require splicing.

e. Prokaryotic and Eukaryotic Translation

- The ribosome is the central player in translation, serving the essential

function of translating mRNA into proteins in prokaryotic cells. In contrast,
eukaryotic cells utilize the ribosome for the process of mRNA-to-protein
translation.
- In prokaryotic translation, a single initiation factor aids in ribosome
binding to mRNA, simplifying the initiation process. Eukaryotic
translation, on the other hand, employs a more complex system with
multiple initiation factors, contributing to the intricate regulation of
ribosome-mRNA interactions.
- Prokaryotic translation streamlines the elongation phase by relying on a
single elongation factor responsible for both mRNA movement and amino
acid addition. In contrast, eukaryotic translation introduces a higher level
of complexity, employing several distinct elongation factors to oversee
these intricate processes.
- In prokaryotic translation, the termination phase is straightforward, as a
single termination factor signals the end of translation and the release of
the polypeptide chain. In eukaryotic translation, this process is more
intricate, involving several different termination factors to manage the
termination and release of the polypeptide chain.

2. Why is post-transcriptional modification of pre-mRNA important for eukaryotic

organisms?
Post-transcriptional modification of pre-mRNA is a crucial process in
eukaryotic organisms, serving several vital functions. It involves the removal of
introns through splicing, which is essential for generating mature mRNA
molecules capable of translation. Additionally, post-transcriptional modifications
add a protective 5' cap and a 3' poly(A) tail to mRNA, safeguarding it from
degradation and enabling ribosome binding and nuclear-cytoplasmic transport.

These modifications also play a pivotal role in gene expression regulation.

Splicing can produce various protein isoforms, while the 5' cap and 3' poly(A) tail
can influence mRNA stability and translation efficiency. Furthermore, this process
is indispensable for specific biological phenomena.

They enable extensive antibody diversity through alternative splicing,

equipping the immune system to combat a wide range of infections. These
modifications also play a critical role in regulating cell growth, differentiation, and
apoptosis, with microRNAs (miRNAs) influencing mRNA translation. Conversely,
mutations in genes tied to post-transcriptional modifications, such as BRCA1 and
BRCA2, can heighten the risk of conditions like breast cancer.

In conclusion, post-transcriptional modification of pre-mRNA is a

fundamental process governing gene expression and many essential cellular
functions in eukaryotic organisms. It underpins the remarkable diversity and
adaptability of the proteome, influences the regulation of cell growth and
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differentiation, and is associated with the development of various diseases,

including cancer.

3. What are spliceosomes? What vital functions do they perform? And what are the
consequences if, in the performance of their functions, they go awry?

Spliceosomes are essential for the production of functional proteins in

eukaryotic organisms. They remove introns from pre-mRNA transcripts,
producing mature mRNA molecules that can be translated into proteins. Mutations
in genes that encode proteins involved in splicing can lead to a variety of diseases,
including cancer. If spliceosomes malfunction, introns may not be removed
properly, resulting in the production of non-functional or truncated proteins.
Additionally, spliceosome malfunction can lead to the production of multiple
protein isoforms, which can have different functions and some of which may be
harmful.

4. Explain Thomas Cech’s discovery of mRNA as a ribozyme. Explain how it ‘breaks

biological rules’.

Thomas Cech's groundbreaking discovery unveiled the catalytic potential

of RNA, challenging established beliefs. It demonstrated that RNA could serve as
both a carrier of genetic information and an active catalyst, defying the
conventional notion that only proteins could function as catalysts. Moreover,
Cech's findings blurred the rigid boundaries of the central dogma of molecular
biology, which primarily focused on the DNA-to-RNA-to-protein flow of genetic
information. Cech's work primarily revolved around the tetrahymena ribozyme, a
small RNA molecule capable of self-catalysis, even in the absence of proteins,
which defied prior assumptions about enzymatic activity. This groundbreaking
revelation revolutionized our comprehension of biology, leading to the exploration
of numerous other ribozymes and their vital roles in various biological processes,
significantly influencing gene expression, protein synthesis, and RNA processing,
while also catalyzing advancements in research and biotechnology.