Transcription and translation Flashcards
Where does transcription occur?
-In the nucleus
What is transcription?
-The first stage in protein synthesis where an mRNA copy is made of a gene of interest from DNA
What chromatin can be transcribed?
-Euchromatin
Why does transcription require ATP?
-In order to start the process once all the transcription factors and enzymes have made the transcriptional complex
What are the three stages of transcription?
- Initiation
- Elongation
- Termination
What is a promoter?
-A recognition sequence of nucleotides on a length of DNA which would initiate transcription when recognised by appropriate transcription factors
Can genes only be in one direction on DNA?
-Can be in both directions as the double strands of the alpha helix are antiparallel
What is the benefit of having a number of promoters?
-Different genes have different promoter regions in order for different genes to be activated in response to different stimuli
What is the ubiquitous region in all promoters of cells?
-TATAAA box
At what position is the TATAAA box located in eukaryotes?
–30
How does the TATAAA box initiate transcription?
- Transcription factors recognise and bind to the TATAAA box
- This binding to the promoter region causes the recruitment of RNApolymerase
- RNApolymerase then moves along the length of DNA until it reaches the start codon
What is position +1 in transcription?
-The location at which RNApolymerase reaches the start codon
Anything upstream from +1 is…
Anthing downstream from +1 is…
- Negative
- Positive
What is the benefit of having regulatory sequences upstream of the TATAAA box?
-Allows other transcription factors to regulate gene expression
How can regulatory sequences 1000s bp away from the TATAAA box influence transcription?
- 3D folding of the protein allows interaction, either through direct interaction or between the regulatory region and a mediator
- Enhancers, inhibitors, activators etc
How does elongation occur in transcription?
- DNA helix unwinds itself
- RNApol reads the 3’->5’ strand in order to produce an mRNA copy of the gene of interest in a 5’->3’ strand
- Phosphodiester bonds join ribonucleotides to make a copy of the gene
What is an ORF in mRNA?
-The section of mRNA which codes for protein
How does termination of transcription occur?
- RNApol reaches the end of the coding region of the gene and dissociates from DNA
- Specific exonucleases recognises sequence in capped nascent mRNA and cuts free from RNApol
What is an nuclease, endonuclease and exonuclease?
- Nuclease-> enzyme which degrade nucleic acids
- Endonuclease->cuts within the polynucleotide; can be specific recognising sequences or non-specific making random cuts
- Exonuclease-> degrades polynucleotides from the 5’ or 3’ end
What are the three processes which result in RNA maturation?
- Capping
- Tailing
- Splicing
What is capping?
- The immediate addition of 5’ cap when mRNA synthesis has begun
- Methylated guanine 5’-5’ phospho linkage
What is the function of the 5’ cap?
- Stops degradation of the polynucleotide
- Plays a role in translation
What is tailing?
-The addition of multiple adenines by polyA polymerase
What is the function of tailing?
-A very long polyA tail (approx 200n) protects the 3’ end of mRNA from 3’specific exonucleases
What is splicing?
-Introns in mRNA removed and exons ligased
Is splicing random?
- No
- It is sequence dependant; introns have 3 specific sequence sites, one at either end and one in the middle
What protein complex carries out splicing?
-A splisosome made of many proteins
Can splicing defects cause disease?
-Yes they can cause unstable or unreadable mRNA
What is a triplet codon?
-A three letter segment which denotes a protein
5’->3’ translation produces a polypeptide in what direction?
-N’->C’
What is the start codon?
-AUG (methionine)
How many termination codons are there?
-3
Why is the genetic code described as degenerate?
-More than one codon can code for one a’a
Why do mutations in the 3rd position of a codon often have no effect?
- Due to the genetic code being degenerate
- Substitutions in the 3rd position of another nucleotide often codes for the same a’a
What are the three main types of RNA?
- mRNA
- rRNA
- tRNA
Why can mRNA copies of the same gene produce different proteins?
-Due to alternate splicing
Why can some mRNA molecules produce proteins with different half lives?
-Different extents of polyadenylation
What is mRNA?
-A copy of the gene of interest
What is rRNA?
-RNA which encodes for ribosomal subunits
Why are there only 4 types of rRNA?
-Only 2 ribosomal subunits
What is the structure of tRNA?
- A single strand of RNA which forms stem loops
- The secondary structure forms the anticodon loop
- Naturally uncharged and free
How do a’as bind to tRNA?
-A bond between the 3’-OH and the COOH of the a’a
What is a tRNA molecule with a bound a’a called?
-AminoacyltRNA
What is the anticodon loop?
-A loop of DNA which contains a complementary codon to the a’a it will bind
How does the anticodon bind an a’a?
-Recognition of the a’a codon is in a antiparallel fashion
What is nucleotide I and why does tRNA have it?
- Inosine - a derivative of purine
- Due to the genetic code being degenerative; I can bind to any of the 4 nucleotides
Why does I introduce a wobble base, and what does it allow?
- I at the 5’ end of the anticodon introduces more space at the 3’ end of the anticodon
- This allows unusual base pairing between nucleotides; if the wobble is large enough G can bind to U
Where does translation occur?
-In the cytoplasm
What is translation?
-The conversion of the mature mRNA molecule into a fully functioning protein
How are tRNA molecules activated?
-Activation occurs by the transfer of a molecule of ATP and the a’a onto the tRNA molecule catalysed by tRNA synthetase
Describe initiation of translation
- Cap binding proteins recognise the mature mRNA molecule
- Met, tRNA and the 40s subunit bind to the mRNA
- tRNA becomes activated as the complex moves downstream and recognises the start codon
- 60s ribosomal subunit and translational factors bind
- GTP, initiation factors and cap binding proteins released
- Fully functional ribosome ready for translation
Describe elongation of translation
- Ribosome has P and A site
- P site occupied by met-tRNA and A site free
- Binding of next aminoacyl-tRNA complementary to next codon
- Peptide bond formed between P and A site via peptidyltransferase
- Ribosome moves along; second a’a now occupies P site and cycle begins again
Describe termination of translation
- Stop codon recognised by uncharged tRNA-lookalike molecule
- tRNA-lookalike physically moves the releasing factor
- Releases new polypeptide chain and mRNA molecule
- tRNA binds -OH group to terminate growth
Can only one ribosome bind to one mRNA molecule?
-No, there can be many translating the same length of mRNA
List the differences of bacterial translation
- Simple promoter regions
- Different transcription/translation factors
- Couples transcription-translation
- Different ribosomal subunits
- Different transcription initiation mechanism
- One RNApol/mRNA
- Different RNApol
How can the differences in bacterial transcription/translation be used as an advantage?
-Allows drug targeting of bacterial cells without damage to mammalian cells during infection
What mutations can occur, causing disease, in areas outside the coding region?
- Promoter/enhancer regions
- Splicing
- Termination regions
- Ribosome binding regions
