0
Q
What is needed in transcription?
A
- Enzyme: RNA polymerase
- Activated substrates: NTPs
- Template: DNA
- 3 stage process: initiation; elongation; termination
1
Q
What is needed in DNA replication?
A
- Enzyme: DNA polymerase
- Activated substrates: dNTPs
- Template: DNA
- 3 stage process: initiation; elongation; termination
2
Q
What is needed in translation?
A
- Enzyme: ribosome
- Activated substrates: amino acids
- Template: mRNA
- 3 stage process: initiation; elongation; termination
3
Q
Describe the 3 stages of initiation, elongation and termination in DNA replication
A
- Initiation: recognition of origin of replication; initiation proteins; DNA polymerase
- Elongation: 5’ to 3’ chain growth
- Termination: replication forks meet
4
Q
Describe the 3 stage process of initiation, elongation and termination in transcription
A
- Initiation: promoter recognition; transcription initiation factors; RNA polymerase
- Elongation: 5’ to 3’ chain growth
- Termination: sequence dependent
5
Q
What happens during initiation of transcription?
A
- Transcription factors bind to the initiation code - 5’ TATA 3’ ie promoter region are upstream to the Open reading frame
- Binding of transcription factors to DNA attracts RNA a polymerase to start mRNA production
- RNA polymerase separates the DNA strands so the RNA nucleotides can bind along the template strand
6
Q
What happens during the elongation stage of transcription?
A
- RNA polymerase travels along the template strand reading it from 3’ to 5’
- Picks up base pairs and copies them onto a complementary RNA sequence forming an mRNA sequence
- The mRNA transcript has nucleoside triphosphates (NTPs) added to the 3’ end (so is made from 5’ to 3’)
7
Q
What is a promoter sequence?
A
- Area of gene upstream of the open reading frame that regulates transcription
- Includes sequences for the binding of transcription factors, RNA polymerase and regulatory factors
8
Q
What happens during the termination stage of transcription?
A
- A methyl-guanine ‘cap’ is added to the 5’ end by a 5’ - 5’ triphosphate linkage
- Stabilises the mRNA
- A stop codon at the 3’ end activates the cleavage of the mRNA (AAUAA)
- 3’ end is then polyadenylated - called tailing (polyAtail)
- Both protect against degradation
9
Q
What happens during the splicing stage of transcription?
A
- Pre-mRNA (with introns and exons) is converted to mature mRNA (exons only)
- Introns are removed by endonucleases (breaks within the polynucleotide) and exonucleases (degrades polynucleotide from 5’ or 3’ end)
10
Q
Describe mRNA
A
- Made by RNA polymerase II
- ~2%
- 100,000 of kinds
- A few copies of each present
11
Q
Describe rRNA
A
- Made by RNA polymerase I
- > 80%
- Few kinds
- Many copies of each
- Eukaryotes: 80s (60s and 40s subunits)
- Prokaryotes: 70s (50s and 30s subunits)
12
Q
Describe tRNA
A
- Made by RNA polymerase III
- ~15%
- ~100 kinds
- Very many copies of each
- Is uncharged without a bound amino acid, becomes charged and is known as an aminoacyl-tRNA when an amino acid is bound
- Has an anticodon which is completely to the codon on mRNA
- Is single stranded RNA molecule that form a clover shape by hydrogen bonding between complementary anti-parallel bases
- Goes from 5’ to 3’ - 3’ end bonds to amino acid
13
Q
What are the features of the genetic code?
A
- Changes from a 4 letter ‘DNA language’ to a 20 letter ‘protein language
- Triplet code
- Degenerate
- Non-overlapping and ‘comma-less’ (no gaps)
- Adaptor molecule is tRNA
14
Q
What is the initiation triplet code?
A
- AUG (codes for methionine - is at the beginning of every protein until it is spliced off)
15
Q
What are the termination triplet codes?
A
- UAA
- UAG
- UGA
- Are all stop codons
16
Q
What is the ‘wobble’ position?
A
- 5’ base of anticodon and 3’ base of codon
- Allows a single tRNA species to recognise more that one codon
- Contains Ionoside nucleotide which is aspecific
17
Q
What happens during the initiation stage of translation?
A
- The 40s subunit of a rRNA binds with Met-tRNA to the 5’ cap of the mRNA
- Starting codon 5’ AUG is recognised and is specific to Methionine only (has the anticodon 5’ CAU
- 60s subunit binds
18
Q
What happens during the elongation stage of translation?
A
- rRNA has two sites for tRNA to bind: P site (holds peptide chain); A site (accepting tRNA)
- Met-tRNA occupies the P site and then another aminoacyl-tRNA enters the ribosome to occupy the A site (requires GTP)
- Methionine forms a peptide bind with the adjacent aminoacyl-tRNA (requires peptidyl transferase) making the tRNA in the P site now uncharged
- This tRNA leaves the ribosome, which moves along the mRNA to the next codon leaving the A site unoccupied again (translocation)
- mRNA is read from 5’ to 3’, and the polypeptide chain grows from amino (N-) to carboxy (C-) terminus
19
Q
What happens during the termination stage of translation?
A
- Requires a stop codon to be read on the mRNA
- Are no tRNA molecules with a complementary anticodon that can bind
- Peptide and tRNA are hydrolysed to release the protein into the cytoplasm
20
Q
Define the term gene
A
- A unit of hereditary
- A transcription unit (ie a length of DNA on a chromosome that contains the code for a protein (or RNA) as well as sequences necessary for its expression, such as promoter and terminator sequences and introns)
21
Q
What are the reactions that occur in the the maturation of mRNA A?
A
- Capping
- Polyadenylation
- Splicing
22
Q
What do bacteria in regards to gene expression?
A
- Simpler promoters
- Different transcription factors
- Single RNA polymerase
- Coupled transcription-translation
- No post-transcriptional processing
- Short-lived mRNAs
- Simpler ribosomes (can be exploited by drugs targeting 30s subunit so it only affects bacteria)
- Distinctive translation inhibition mechanism
- Different translation factors
23
Q
How can the effects of various mutations in a gene be predicted?
A
- Severity of mutation depends on the amount of difference in a protein
- If an amino acid is swapped for one with similar properties there will not be a large effect
- Effects may be profound however if amino acids have different properties
- Premature or delayed stop codons cause truncated/elongated proteins that may not function correctly or at all
24
How can mutations outside the coding region affect gene expression?
- Mutations to promoter regions where transcription factors bind can activate or deactivate gene expression
MGD
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