DNA Transcription And Translation Flashcards
How does amplification happen in protein synthesis?
1-2 copies of gene in a cell
Several copies of same RNA can be made
Several protein molecules are then made from each RNA
Why is the 3D shape of RNA hugely varied?
Because RNA is usually single stranded, forming a wide variety of shapes
What difference is there between RNA polymerase and DNA polymerase?
RNA polymerase can initiate RNA chain growth without a primer
What is the non-coding strand also known as?
The template strand
What is transcription the process of?
DNA —> RNA
What do mRNAs do?
Code for proteins
What do rRNAs do?
Form the core of the ribosome’s structure and catalyse protein synthesis
What do miRNAs do?
Regulate gene expression
What do tRNAs do?
Serve as adaptors between mRNA and amino acids during protein synthesis
What important promoter region is present in bacteria and eukaryotes?
TATA box
What’s the reason for having 2 promoter regions in bacterial DNA?
Second promoter region - 35BP away from start site - gives directionality of synthesis
In which strand does the promoter sequence lie in?
Promoter sequence sits in the coding strand
RNA polymerase then reads the opposite - template - strand to create RNA chain homologous in sequence to coding strand
What’s the key difference between eukaryotic and prokaryotic systems with respect to RNA?
There are multiple RNA polymerase in eukaryotic systems
Bacteria have 1 RNA polymerase
Eukaryotes have 3
What genes do the 3 different RNA polymerases transcribe in eukaryotes?
RNA Poly l - most rRNA genes
RNA Poly ll - all protein-coding genes, miRNA genes, plus genes for other non-coding RNAs
RNA lll - tRNA genes
What RNA polymerase variant synthesises mRNA?
RNA polymerase ll
What further complexes transcription in eukaryotes?
Larger number of accessory proteins needed for RNA polymerase
Much more elaborate control of transcription mechanism - JUNK DNA
Chromatin packing consideration is needed
Where does translation occur?
In cytoplasm
Where does transcription occur?
In nucleus
How far up stream is the TATA box in eukaryotes/prokaryotes?
Eu - 25 nucelotides
Pro - 10 nucelotides
Why are phosphates added to RNA polymerase ll in transcription?
Phosphates are added to RNA-Pll to disengage from transcription factors so transcription can begin
What’s the reason for the 90˚ kink that happens at TATA box?
Partial unwind of DNA which leads to recognition
How is a RNA molecule post-transcriptionally processed to form mRNA?
Capping factors are attached to 5’ end early in synthesis - after 25 nucleotides
Polyadenylation factors added to 3’ end
Why is mRNA capped and polyadenylated?
Increases stability
Aid export from nucleus to cytoplasm
Protein synthesis machinery can check mRNA to ensure it is fully intact - both sites must be present
What splices introns out?
Spliceosome - key component is some catalytic RNA
When does splicing begin?
Splicing occurs after capping and whilst mRNA continues to grow.
When does splicing cease?
Either just before or just after polyadenylation occurs
Why is splicing not a wasteful process?
Because mRNA transcripts can be spliced in different ways to produce different proteins
(Alternative splicing)
What does alternative splicing explain?
That we have 100,000 proteins but only 24,000 genes
What do transcription regulators control?
They control how often transcription is initiated
Where do transcription regulating proteins bind?
Usually at the major groove
How many interactions are formed between TRPs and the DNA sequence?
10 - 20
(Strong net force)
What are synthetic polyamides and what is their purpose?
They are molecules capable of recognising and binding to specific base pairs
Anticancer agents
Where are synthetic polyamides capable of binding?
Capable of minor groove binding
What functional group are present in synthetic polyamides?
Heterocyclic rings
What is the MoA of synthetic polyamides?
Molecule attaches to DNA like a clamp - each arm binds to one DNA strand each
Bind where transcription factor usually would
What are the potential uses of short oligonucleotides?
Antiviral and anticancer potential
What are the benefits of hurt short oligonucleotides?
Highly specific - low doses result in fewer side effects than conventional protein inhibition
What are the disadvantages of short oligonucleotides?
Section of mRNA chosen must be exposed - poor absorption and susceptible to metabolism
What do short oligonucleotides do?
They are used to block coded messages carried by mRNA
What is redundant coding?
Where multiple codons code for the same amino acid
What are the 3 stop codons?
UAA
UAG
UGA
What is always the start codon?
AUG - methionine
Since all proteins start with AUG, do all proteins start with methionine?
No, Methionine is post-translationally removed if necessary
Where do tRNAs vary at?
At the anticodon and are attached to an amino acid
How long are tRNAs roughly?
≈ 80 nucleotides long
How many segments do tRNAs consist of?
4 short segments form double helical sections
Clover leaf shape
What are the 2 unusual bases which tRNAs contain?
Dihydrouridine (D)
Pseudouridine (Ψ)
When are D and Ψ created?
They are created by chemical modification post-transcriptionally
Why is the anticodon region important in tRNAs?
Because it is the region that binds to mRNA
How does redundant coding relate to tRNA?
Some aa have more than one tRNA it can attach to
Some tRNAs are made so they require accurate base pairing only at first two positions
How are tRNAs charged with amino acids?
Each amino acid is linked to it own aminoacyl-tRNA synthase protein.
The synthase recognises the correct tRNA for its amino acid.
This process is called charging and forms high energy bonds
What happens after charging?
The tRNA anticodon forms base pairs with the codon on the mRNA
What is the combined mass of the large and small subunit of ribosomes?
10^6 daltons
Standard protein is ≈ 30,000 daltons
What way do ribosomes read mRNA?
5’ —> 3’ manner - synthesised from N-terminus to C-terminus
Uses 5’ - cap as signal
Why is charging important?
Because tRNA can’t bind mRNA without being charged first by correct amino acid
How many RNA molecules are in ribosomes?
4
What’s the purpose of the small ribosomal subunit?
Matches the tRNA to the codon in mRNA
What’s the purpose of the large ribosomal subunit?
Catalyses the formation of the peptide bonds during polypeptide chain growth
What’s the rate of decoding in eukaryotes?
2 amino acids per second
What’re the 3 site in ribosomes?
A site - Aminoacyl-tRNA site
P site - Peptidyl-tRNA site
E site - Exit site
What’s important to remember about the 3 factors of ribosomes?
Only 2 sites are ever filled at anytime one time
What’s the benefit of polyribosomoes?
Many ribosomes can read the same mRNA strand at once
Amplified
What do initiation factors do?
They help the small subunit locate the starting point of mRNA
5’ cap is recognised by small subunit
What happens once 5’ cap has been recognised?
Small subunit and initiation factors move along mRNA until start codon is reached
What happens once AUG has been reached?
Translation initiation factors dissociate
Large subunit binds - forms around mRNA and tRNA
Translation begins
What is the first step once translation begins?
The correct charged tRNA enters in the A site and binds to the mRNA for long enough for the mechanism to work
Large subunit catalyses the first peptide bond
Trial and error - slow
What happens once first peptide bond is formed?
New peptide bond is in A site which is then translocated into the P site - middle
The process continues
How does the growing peptide chain move from the A site to the P site?
The large subunit translocates across to move peptide chain into the P site
The small subunit also then translocate to get back into standard position
The previous tRNA is now in the E site and is ejected
What happens in the final phase of protein synthesis?
The binding of release factor to an A-site bearing a stop codon terminates translation
What does binding of the release factor do?
It alters the activity of the peptidyl transferase in the ribosome, catalysing the addition of water instead of an amino acid
This frees up the carboxyl-end of the peptide
What is tetrecycline’s specific effect?
Blocks binding of aminoacyl-tRNA to A site of ribosome
What’re the specific effects of streptomycin?
Prevents the transition from initiation complex to chain elongation
Also causes miscoding
What’s chloramphenicol’s specific effect?
Blocks the peptidyl transferase reaction on ribosomes
What’s the specific effects of cycloheximide?
Blocks translocation reaction on ribosomes
What’s the specific effect of rifamycin?
Blocks initiation of transcription by binding to RNA polymerase
