Preceptor Session 6
October 3rd, 2024
BIO 300
Chapter 7
From DNA to Protein
The central dogma
DNA → RNA : Transcription
RNA → Protein : Translation
Important because it creates the proteins
that are needed for our cells to function
Transcription
DNA to RNA
Looking at structure of RNA
Backbone: phosphate, sugar
- Sugar: ribose
Talked about in 5’ → 3’ direction
Has a uracil base instead of thymine
Single stranded (not always tho)
Unstable and temporary
- Unstable because it’s only single stranded
- Temporary because that’s who RNA is
Uracil and thymine are like twins
Uracil and thymine are structurally similar,
so..
- Uracil can bind to adenine
- Uracil and adenine have 2 hydrogen
bonds between them
There are many types of RNA
Messenger RNA (mRNA) : codes for
proteins
Transfer RNA (tRNA) : links amino acids to
mRNA during protein synthesis
Ribosomal RNA (rRNA) : form the core of
ribosome’s structure and catalyze protein
synthesis
Transcription is similar to DNA replication
Both open up DNA to expose its bases
Both use 1 strand of DNA as a template for
complementary base pairing
- Either side of DNA can be used for
transcription
- U binding to A instead of T
Both involve polymerases
- RNA polymerase instead of DNA
polymerase
Both synthesize in the 5’ to 3’ direction
RNA polymerase vs DNA polymerase
RNA polymerase adds ribonucleotides to the
3’ end, not deoxyribonucleotides
RNA polymerase does NOT need a primer to
get started like DNA polymerase
RNA polymerase makes more mistakes
- It doesn't have a proofreading site like DNA
polymerase
- It’s okay that it makes mistakes because
RNA is temporary
Start and stop recognition sequences
Start: promoter region
- This is where RNA polymerase binds to
start transcription
Stop: terminator region
- This is where RNA polymerase stops
transcription
Bacterial transcription
1) Sigma factor and bacterial RNA polymerase Sigma factors: proteins that bind to bacterial
bind together RNA polymerase and recognizes promoter
regions
2) They take a stroll on the DNA strand until it
recognizes a promoter region
3) Sigma leaves, then the polymerase latches
to DNA strand and begins RNA synthesis
4) Synthesis stops once polymerase hits a
terminator region
5) Both polymerase and RNA chain release
One bacterial mRNA can code multiple
proteins
Bacteria can express a bunch of different
genes on the same piece of mRNA
- Multiple proteins can be made from 1
mRNA
Eukaryotes CANNOT do this because
mRNA here only has information for 1
protein
Eukaryotic vs Bacterial transcription
Eukaryotes have several types of RNA
polymerases
- Only need to know about RNA polymerase
2
- Does the transcribing
Eukaryotic RNA polymerase require
transcription factors
Eukaryotic transcription is more elaborate
and complex
General transcription factors
Proteins that determine which genes to
express and when
Functions
- The first proteins that bind to the promoter
area
- Position and launches RNA polymerase
- Pulls apart DNA to expose bases
Eukaryotic transcription
1) Transcription Factor 2 D (TF2D) binds to TATA box: short DNA sequence found in
TATA box promoter region. A-T rich region
2) Other transcription factors pile on - A-T rich because it’s easier to break due to
less bonds
3) RNA polymerase 2 gets recruited
4) Transcription Factor 2 H (TF2H) opens up
DNA strands with energy from ATP
hydrolysis
5) TF2H phosphorylates tail of RNA
polymerase 2
6) RNA polymerase 2 is now on and starts
transcription
Transcription initiation complex
It’s a collection of general transcription factors
and RNA polymerase 2
The order of transcription factors piling on
differs by the promoter
Phosphatase causes the release of RNA
polymerase 2 from DNA by
dephosphorylating the tail
These are points of regulation!
RNA polymerase can transcribe a gene many
times simultaneously
Once an RNA polymerase starts
transcribing and the promoter region is
free, another one comes in, then another,
then there’s multiple transcribing
RNA processing
Our RNA has to be processed in order to
move it from DNA to the cytoplasm for
translation
- Capping
- Tailing
These are happening due to enzymes that are
part of the RNA polymerase
Processing occurs when synthesized RNA
comes out of RNA polymerase
Capping and Tailing
These have to be done in order for our RNA to
move from DNA to the cytoplasm
Capping (5’ Cap)
- Big functional structure that caps the 5’
end
- Bound by triphosphate
Tailing (Poly A Tail)
- Bunch of adenines at the end
Eukaryotic vs bacteria mRNA
Bacteria
- The whole gene is a coding region
Eukaryotic
- The gene has both coding and non coding
regions
- Coding: Exons
- Non coding: Introns
- These have to be removed for gene
expression to occur
The spliceosome
This is a collection of RNA proteins that
carries out splicing
- Splicing: Introns get cut out and the exons
get joined together
Small nuclear ribonucleoprotein particles
(snRNPs) form the core of the spliceosome
and carries out the splicing
Alternative splicing allows protein diversity
This is splicing exons from the same gene
into different combinations to make different
proteins
- Point of regulation
- Increases the coding potential of the
genome
Mature mRNAs go to cytoplasm
Mature mRNA
- mRNA + capped + tailed + spliced
Once it's found to be mature, it leaves the
nucleus through nuclear pores
Eukary. & prokary. transcription in picture form
Review questions
1) What base does RNA have that DNA 6) What proteins bind to the promoter area
doesn’t? What is its complementary base? first and determine which genes are being
expressed?
2) What kind of RNA codes for proteins?
7) What is the TATA box?
3) Why does RNA polymerase make more
mistakes than DNA polymerase? Why is this 8) What is the difference between a
okay? eukaryotic and bacterial gene?
4) The promoter region is where 9) What 3 things must happen before RNA
transcription ______, while the terminator can go to the cytoplasm?
region is where transcription ______.
10) What protein particles form the core of
5) Are eukaryotes or prokaryotes able to the spliceosome and carries out the
code different proteins on one mRNA? splicing?
Translation
RNA to Proteins
The protein code
Codons: 3 consecutive nucleotides that
specify an amino acid
Start codon: AUG (methionine)
Stop codons: do not code for an amino
acid and stops translation
Be careful reading the code
When reading mRNA, it has to be read in
the correct 3 letter reading frame
If not, it could completely change the
amino acid sequence
Ribosomes
A big protein complex that carries out
translation
Has a large and small subunit
- Both subunits are made from ribosomal
proteins and rRNAs
If the ribosome is not actively translating, the
large and small subunit are NOT together
Each ribosome binds 1 mRNA and 3 tRNAs
When the two subunits come together, we
get 3 major active sites
- A site: Where new amino acids get added
to growing polypeptide chain
- P site: Growing polypeptide chain
- E site: exit
The tRNAs are going to bind to the large
subunit
The mRNA is going to bind to the small
subunit
tRNA
These bring in the amino acids
Has an upside down clover leaf structure
- Bottom leaf has an anticodon
- Anticodon: sequence that binds to a
complementary codon on mRNA
- The top of the stem (3’ end) is where the
amino acid is attached
Cont’d
tRNAs need energy to bring in the amino
acids. So, they use aminoacyl-tRNA
synthetases
- Enzyme that covalently links amino acid to
tRNA
Energy of bond will later link the amino acid
to the polypeptide chain
Start of translation process
1) mRNA binds to small ribosomal subunit
2) The subunit goes along until it finds the
start codon (AUG)
3) Large ribosomal subunit binds to small
subunit
4) tRNA binds to A site and brings in the first
amino acid
5) First peptide bond is now formed
Middle of translation process
1) Ribosome continues down the mRNA and
reads each codon
2) tRNA is brings in amino acids that are
complementary to the codons
3) Amino acids join together and form the
growing polypeptide chain in P site
End of translation process
1) Ribosome keeps doing its thing until it hits
a stop codon
2) Release factors bind to the codon
3) Ribosome will release the polypeptide
chain
4) Ribosomal subunits separate
Translation occurs at the ribosome
Ribosomes are HUGE and can be seen
under an electron microscope
Ribosomes are actively translating when
they are bound to the endoplasmic
reticulum (ER)
Polyribosomes
There can be multiple ribosomes on a
singular mRNA at the same time
This allows the cell to make copies of a
protein very quickly
Antibiotics that block bacterial protein
synthesis
These drugs target gene expression in
bacteria to inhibit them from making
proteins
*Don’t need to know details, just know that
this is a thing
Genes can be expressed at different levels
Cells are going to regulate how much of a
protein they're going to make
- Point of regulation!
They can transcribe a gene a bunch of times
to make a lot of proteins
Or, they don’t transcribe as much because
they don’t need a lot of proteins
Review questions
1) What are codons? What is the start 6) What enzyme gives tRNA energy?
codon?
7) What is the anticodon region of tRNA?
2) Why do you need to be careful when
8) Ribosomes are translating when they are
reading mRNA?
(bound/unbound) to the ER.
3) What protein complex carries out
9) Multiple ribosomes on a singular piece of
translation?
mRNA are called?
4) What are the 3 active sites formed when
10) How can genes be expressed at different
the subunits of ribosome come together?
levels?
5) What kind of RNA brings in the amino
acids during translation?
