Transcription & Translation Flashcards
Outline the ‘central dogma of molecular biology’
Idea proposed by Crick
Process by which the instructions in DNA are converted into a functional product - since DNA and proteins are structurally and chemically very different therefore DNA cannot be made directly to protein and a messenger molecule (RNA strand - diffusible) is needed
(Replication) DNA ->(transcription) RNA -> (translation) protein
Definition of ‘gene’
A defined region/sequence of DNA that produces a diffusible product (RNA) that has some function
Anatomy of prokaryotic genes - 3 regions
- Coding sequence - stretch of DNA over gene that gives rise to protein
- Promoter region - AT rich region where transcription starts (upstream/before coding sequence). It is the DNA segment recognised by RNA polymerase to start transcription
- UTR (untranslated region) - region of the gene that will be transcribed to be part of mRNA but will not be translated so will not be part of the protein
5’UTR vs. 3’UTR
5’ UTR = short (3 - few 100 nucleotides)
3’UTR = long (1000s of nucleotides)
Functions of UTR
Regulatory functions (2 levels)
- Transcription level: part of DNA so involved in when to move from DNA to RNA (transcription)
- Translation level: part of mRNA so involved in when to move from mRNA to protein (translation)
UTR in modifying mRNA in eukaryotic genes + function
2 types of modification - 1 on each side of mRNA
- 5’UTR side: 5’G cap (chemically modified G nucleotide)
- 3’UTR side: Poly-A-tail (long stretch of A nucleotides 20-250)
Function - both 5’G cap and poly-A-tail prevents mRNA degradation from enzymes
what is the 3’UTR part of
Exon 4 - the last exon
what is 5’UTR part of
Exon 1
Functions of 5’G cap (4)
- Prevents mRNA degradation
- Regulates translation by providing a ribosome recognition and binding site
- regulation of nuclear export
- promote intron excision
Functions Poly-A-tail
- Prevents mRNA degradation
- regulates translation
- regulates nuclear export
Pre-mature mRNA vs. mature mRNA
Pre-mature mRNA has both intron and exon regions
Mature mRNA has intron regions cut out
Important points for both prokaryotic and eukaryotic genes regarding non-coding regions (4)
- contain non-coding DNA regions upstream and downstream of the coding sequence (eukaryotes also have introns)
- the non-coding DNA regions can be transcribed but are not translated
- These non-coding DNA elements are involved in regulating gene expression
- changes/mutations in these non-coding gene sequences may ‘disrupt’ normal gene expression
location of translation - prokaryotes vs. eukaryotes
prokaryotes - in cytoplasm
eukaryotes - in nucleus
What are the 3 stages of transcription (same for translation)
Initiation, elongation, termination
Outline initiation - the first step of transcription
- Transcription factors bind to promotor region (TATA box) to signal RNA polymerase II to bind to this particular region
- RNA pol II binds, forming a transcriptional initiation complex together with the transcription factors
- RNA pol II unwind the DNA, the 2 DNA strands separate and RNA pol II starts transcription (RNA synthesis) without the need of a primer
Outline elongation - the second step on transcription
RNA pol II uses the template strand which runs in the 3’ - 5’ direction, as a template and inserts complementary RNA nucleotides in the 5’ - 3’ direction (A-U, C-G)
1 DNA strand can be transcribed by many RNA polymerases at the same time
Outline termination - the last step in transcription and state difference between prokaryotes and eukaryotes
RNA pol II encounters a particular sequence that code for termination and will disengage from the DNA template
In prokaryotes - the RNA strand will be produced and i ready for translation
In eukaryotes - the pre-mature mRNA strand (with introns) is produced and will undergo splicing to form mature mRNA
Name the 2 ways that eukaryotic cell can produce multiple different proteins from one gene
Alternative splicing and post-translational modifications
Alternative splicing
Exclusion of particular exons during pre-mRNA processing
Post-translational modifications (2)
- Polypeptide processing = exclusion of particular regions of the translated (but still immature) protein chain
- Chemical modifications = attachment of biochemical functional groups to the translated (but still immature) protein which will affect the function and activity of the protein
Another term for coding strand (5’ - 3’)
sense strand
Another term for non-coding strand (3’ - 5’)
Anti-sense strand
RNA polymerase (3 features)
- catalyst in transcription by forming phosphodiester bonds between ribonucleotides
- Has primase enzyme within itself (internal 3’ OH group) that it can use to start RNA synthesis
- also has helicase activity
What is transcript
Newly synthesised RNA
Key features of the genetic code (7)
- There are 20 amino acids but only 4 nitrogenic bases
- Every 3 bases code for an amino acid -> triplet code (triplet codon hypothesis)
- This provides 64 possibilities (4^3)
- 61 out of 64 codons specify an amino acid
- Most amino acids have more than 1 codon (redundancy)
- 3 STOP codons (UAA, UAG, UGA)
- 1 START codon (AUG)
What are the 3 STOP codons
UAA, UAG, UGA
What is the START codon and what else does it specify
AUG, this codon also specifies for methionine (Met) which is the first amino acid that is translated
Adaptor hypothesis
The amino acid is carried to the template (mRNA) by an adaptor molecule (tRNA), and that the adaptor is the part which actually fits onto the RNA
codon
triplet of bases on the mRNA that encodes for one amino acid
anticodon
triplet of bases on the tRNA complementary tot he codon region of the mRNA
Key features of tRNA (the ‘adaptor’ molecule) - (7)
- a single-strand of RNA
- at least 1 tRNA for each amino acid
- 70-80 nucleotides in length
- 3’ end = amino acid attachment site (binding region)
- Anticodon = complementary to mRNA anticodon (interaction region)
- 3D structure = upside down L
- can contain I (Inosine)
Importance of the upside down L structure of tRNA
Allows the tRNA to fit into the pockets of the ribosome which is the machinery that will carry out translation
Total number of tRNA and explain how it is possible given that there are 64 codons
32
This is possible due to the 5’ end containing I (Inosine) which is a base that can bind to A, U or C
This allows for one anticodon to bind to more than 1 condon
Outline the process of ‘charging’ tRNA (4)
The process by which an amino acid binds to a tRNA molecule
- Enzyme aminoacyl-tRNA synthase recognises a specific amino acid and its corresponding tRNA molecule and has a specific site for the amino acid and anther specific site for the corresponding tRNA
- Amino acid and corresponding tRNA binds to specific sites on aminoacyl-tRNA synthase
- The enzyme will catalyse a chemical reaction and form a bond between the amino acid and the tRNA using ATP
- Aminoacyl tRNA released (charged tRNA)
Role of ribosomes in translation
site of protein synthesis using mRNA as a set of instructions
Ribosome - prokaryote vs. eukaryote
same anatomy
Anatomy of ribosome (4)
- contain both rRNA (ribosomal RNA) and proteins
- contains small and large subunits which are separate unless performing translation
- small ribosomal subunit = where the mRNA binds
- large subunit (4 regions)
State the 4 regions of the large ribosomal subunit and the role of each
A site (Aminoacyl - tRNA binding site) - where the charged tRNA carrying an amino acid will enter P site (Peptidyl-tRNA binding site) - where the tRNA carrying the growing polypeptide will enter E site (exit site) - where an empty tRNA (tRNA which has transferred its amino acid onto the growing chain of amino acids) will exit Exit tunnel (located above T site) - channel through which the growing polypeptide chain will exit
Outline initiation - the first step of translation (4)
- The small ribosomal subunits finds the initiation AUG codon on the mRNA
- The AUG codon is positioned in the P site of the small ribosomal subunit
- tRNA ‘charged’ with the amino acid Methionine (Met) binds to the P site
- The large ribosomal unit attaches and completes the initiation complex - energy is required
Outline elongation - second step of translation
The ‘charged’ tRNA, with an anticodon complementary to the A site codon, lands in the A site. Then 2 things happen at the same time
1. Peptide transfer - The ribosome will break the bond that binds the amino acid to the tRNA in the P site, transfer the amino acid to the newly arrived amino acid attached to the tRNA in the A site and form a peptide bond between them. Now we have a tRNA with the growing amino acid chain in the A site and an empty/uncharged tRNA in the P site
2. Translocation - When the tRNAs are bound to the mRNA in the P and A sites, the ribosome moves 3 nucleotides (1 codon) down the mRNA. The tRNA with the growing amino acid chain (which was in the A site) will now be in the P site so that the chain of amino acids can exit through the tunnel located above the P site. The ‘uncharged’ tRNA (which was in the P site) will now be in the E site and detaches from its anticodon and is expelled (Energy required)
A new ‘charged’ tRNA with an anticodon complementary to the next A site codon enter the ribosome at the A site and the elongation process repeats itself
Outline the process of termination - last step of transcription
- When the ribosome reaches a STOP codon, a protein called release factor enters the A site
- The release factor uses water to break the bond between the P site tRNA and the lastly added amino acid. This causes the polypeptide chain to detach from its tRNA and the newly made polypeptide (protein) is released
- The small and large ribosomal subunits dissociate from the mRNA and each other (energy is required)
When is energy required in each step of translation
Initiation - energy is needed to form the translation initiation complex
Elongation - energy is needed to detach and expel the ‘uncharged’ tRNA from its anticodon and out through the E site
Termination - energy is required to dissociate the small and large subunit from mRNA and from each other
What is meant by polyribosomes
a single mRNA may be translated by several ribosomes at the same time
Comparison of prokaryotic and eukaryotic translation + transcription
Prokaryotic
- Transcription and translation both occur in cytoplasm
- Transcription and translation are coupled
Eukaryotic
- Transcription in nucleus
- Translation in cytoplasm
- Transcription and translation are NOT coupled
