Card Set Information
What is the name for codons that specify the same amino acid?
Sequence at 3' end of tRNA
...CCA, terminal 3'OH or 2'OH is aminoacylated
tRNAs attached to amino acids by
aminoacyl RNA synthetases
2 step mechanism for aminoacyl tRNA synthetases
: ATP in, pyrophosphate out
: tRNA in, AMP out
aminoacyl tRNA synthetases recognise
1 type of amino acid
all cognate tRNAs
Accuracy depends on which step in activating tRNA? Example?
The second step: IleRS can charge Val with AMP, but hydrolyses valul-aminoacyladenylate when tRNA
Inosine in the wobble position can pair with
U, C or A
Distribution of mass in ribosomes
Sizes of subunits in bacterial ribosomes
Large subunit (peptide transferase)
Sizes of subunits in mammalian ribosomes
Large subunit (peptide transferase)
Sedimentation coefficient dependent on? Units?
How is protein synthesis initiated with regards to ribosome activation
Small subunit binds to mRNA
Large subunit joins it at the initiating AUG codon
Maximum density of ribosomes
What is at the beginning of bacterial mRNA strands and how does this relate to initiating protein synthesis?
: start with formyl-methionine
Formyl rapidly removed, met may be removed more slowly
formyl-Met-X cut by deformylase then aminopeptidase
What is at the beginning of eukaryotic mRNA strands and how does this relate to initiating protein synthesis?
: start with methionine (AUG)
May be removed later depending on the next amino acid
Met-X, bond broken by aminopeptidase
How do bacteria distinguish between initiating AUG codons and normal AUG codons?
: 5'-GGAGG-3' nearer 5' end
Recognised by CCUCC at the end of the
Each cistron in polycistronic mRNA has its own AUG and Shine-Dalgarno sequence
How do eukaryotes distinguish between initiating AUG codons and normal AUG codons?
'Scanning ribosome model'
eIF4F (eukaryotic initiation factor) on 40S subunit binds to 5' cap
scans along RNA towards 3' end (using ATP)
Locks on at AUG, 60S subunit joins
sometimes, different start sites can be used to generate different proteins from one mRNA
Viral method of initiation of translation
'Internal initiation of translation'
first discovered in
IRES (internal ribosomal entry site) in the 5'UTR directs the binding of the ribosome to the mRNA
How to define IRES sequences
dicistronic assay: put IRES between two coding genes; if 2nd one is expressed, then the IRES works
Major IRES drug target
2 sites on ribosomes for binding charged tRNA
: for peptidyl-tRNA
: for aminoacyl-tRNA
How does translation occur?
Charged aa-tRNA binds to A site, using EFTu (Elongation factor protein) (bac) or eEF-1 (euk)
GTP hydrolysis causes the dissociation of EFTu from the ribosome
A peptide bond forms from the peptide held by the tRNA already in the P site eg by
The bond between the P site peptide and the P site tRNA is broken
The ribosome moves 3 nucleotides along the mRNA (
) using EFG (bac) or eEF-2 (euk) and GTP
The peptidyl-tRNA (now with its peptide joined to the peptide next door) moves into the P site, while the deacylated peptide (that WAS in the P site) is kicked out, perhaps through the
Two proteins that use GTP in translation (bacterial and eukaryotic for each)
(euk) bind the aminoacyl-tRNA to the A site
(euk) cause translocation of the ribosome
How is transcription terminated?
If a stop codon (UAA, UAG, UGA) is in the A site,
(release factor) binds, meaning peptidyl transferase binds to polypeptide to H2O -
of the peptidyl-tRNA and therefore release
2 protein synthesis inhibitors
: covalently modifies
- cell death
: ribsome inactiated by removal of a single
base from the eukaryotic 28S rRNA via its glycosidase activity
5 antibiotics that affect protein synthesis
: binds to 30S unit, inhibits initiation, or causes misreading of mRNA at low concentrations
: inhibits binding of aminoacyl-tRNA to the 30S subunit
: inihbit formation of initiation Met
-30S ribosomal subunit-mRNA complex
: inhibits peptidyltransferase activity of 50S subunit (for typhoid and bacterial meningitis)
: blocks progression of nascent peptide by binding to 50S subunit by the peptide exit tunnel
Two types of control of translation
: eg interferon response
: eg ferritin
What is the interferon response?
is an initiation factor that binds Met-tRNA
and GTP and delivers them to the 40S subunit
eIF2's activity is REDUCED by PHOSPHORYLATION
is activated in response to
, which is usually a by-product of viral infection
replication is inhibited
An example of specific translation control
protects the cell from too much iron by binding it
When [Fe] is low,
(iron regulatory protein) binds to
(iron response element), which is in a hairpin in the 5'UTR of the ferritin gene
IRP binding to IRE means the 40S subunit cannot bind to the 5'cap
-> Initiation of ferritin synthesis does not happen when [Fe] is low
What is RNAi?
What are riboswitches? example?
RNA sequences in 5'UTR/introns that bind small molecules to regulate transcription
levels directly control thiamine synthesis
Two ways to degrade proteins
How do lysosomes work?
Non-selective in well-nourished cells
: degrade longlived proteins and organelles all the time
Selective after prolonged fast
Import and degrade proteins with
Lys-Phe-Glu-Arg-Gln (KFERQ) - selectively lost from tissues that atrophy in response to fasting eg liver, kidney, not brain, testes
How do proteasomes work?
Mediate destruction of short lived and ubiquitinated proteins
How is ubiquitin linked to proteins?
(ubiquitin activating enzyme) links
in Ub to
in E1 forming a
(ubiquitin conjugating enzyme) ligates Ub to itself
ligates Ub to the
e-NH2 groups of lysines
in the protein destined for destruction
This is related to Cyclin B?
How are ubiquitins removed?
4 stages of proteasomal destruction
: Ubiquitinated proteins bind to proteins in cao
: ATP dependent process unravels the protein and spits out the Ubs
: The protein is fed into the shaft - the inner two rings are responsible for proteolysis
: protein is entirely degraded into 8 amino acid peptides which are released
3 other types of protein destruction sequences
: 5 specific amino acids at N-terminus
: Pro, GLu, Ser, Thr - phosphorylated on S or T
: seems to be a sequence required for cyclin ubiquitination