Translation Flashcards
What is the first step in protein synthesis?
Translation
-the code contained in a mRNA is translated into a particular series of amino acids to form a polypeptide
What are the mRNAs, tRNAs, and RRNAs synthesized during transcription used for?
To complete translation
What does the protein coding region of an mRNA contain?
A series of nucleotide triplets
What is each Triplett called?
Codon
How many possible combinations of nucleotides are there?
64
How are the nucleotides read?
5’ to 3’
In what direction is the protein synthesized?
From its N-terminus to its C-terminus
Start codon
AUG
-for all protein synthesis and defines the reading frame
Reading frame
The serious of triplets that make the subsequent codons
WHat does AUG code for, besides being the start codon?
Methionine
Are all AUGs start codons?
No, but all start codons are AUG
How many codons does methionine have?
1
Internal codons containing AUG
They are methionine codons, not additional start codons
The first AUG encountered reading 5’ to 3’
Defines the start codon and the subsequent reading frame
Stop codons
- do not encode an amino acid
- UAG, UAA, UGA
___ of the 64 codons define as amino acid
61
-AUG defines an amino acid, but 3 are stop codons and do not code for AA
Codon specificity
Each codon is specific to a certain amino acid
Codons-universal
Genetic code defines the same amino acids in almost all organisms
Codon redundancy
Each amino acid may have more than one codon
How many codons do each amino acid have?
Between 1 and 6
Codon: nonoverlapping and commaless
- code is defined as a continuous series of 3 basses, no overlap and no “punctuation” (no codon is read more than once and no bases are skipped or repeated)
- should start at beginning and read through to the end
Do stop codons code for amino acids?
Nope
START DOES!
What are some examples of single nucleotide changes?
- Silent mutation
- Messenger mutation
- Nonsense mutation
Single nucleotide mutations
Point mutation
Silent mutation
- single nucleotide mutation
- if the DNA sequence is mutated, so that the codon is changed, but still encodes the same amino acid, it is called a silent mutation
UCA to UCU, both encode for serine
Missense mutation
- single nucleotide mutation
- point mutation
- if the point mutation results in a codon that defines a different amino acid, it is called a missense mutation
- can be conservative
UCA to UCU serine to proline
Conservative mutation
-missense mutation that results in an amino acid with similar properties (not as serious, non-polar AA to another non-polar AA)
Nonsense mutations
- single nucleotide mutation
- point mutation
- if the mutation results in a change from an amino acid to a STOP codon
UCA to UAA serine to stop codon
Frameshift mutations
- insertion/deletion is NOT a multiple of 3
- usually result in a premature stop codon and a truncated protein; the closer to the beginning of the protein, generally the more severe the mutation (similar to nonsense)
If the insertion/deletion is not a multiple of 3
Frameshift mutation
If the insertion/deletion is a multiple of 3
You will have an insertion/deletion of amino acids
Splice site mutations
Changes in nucleotides involved in splicing RNA
Splice site mutations could result in any of the following
- deletion of nucleotides from an exon
- leaving nucleotides from an intron in the final mRNA
- completely deleting an exon from the final mRNA
Can be cause by single nucleotide (point) mutations
Trinucleotide repeat expansion
Regions in a gene where a sequence of bases is repeated many times can undergo trinucloetide repeat expansions
-the repeat is amplified significantly
Trinucleotide repeat expansion in the coding region of mRNA (reading frame)
A faulty protein
Trinucleotide repeat expansion in the 5’ or 3’ UTR
Decreased production of the protein due to the effect of the UTRs on translation
-usually underexpressed
Effect on protein: silent mutation
None
Effect on protein: missense
Possible decrease in function; variable effects
Effect on protein: nonsense
Shorter than normal; usually nonfunctional
Effect on protein: frameshift
Usually nonfunctional; often shorter than normal
Effect on protein: splice donor or acceptor
Variable effects ranging from addition or deletion of a few AA to deletion of an entire exon
Effect on protein: triplet repeat expansion
Expansions in coding regions cause protein product to be longer than normal and unstable
Disease often shows anticipation in pedigree
Components required for translation
- amino acids
- tRNA
- aminoacyl-tRNA synthetases
- tRNA charging
- mRNA
- ribosomes
- protein factors
Amino acids in translation
All amino acids encoded on the mRNA must be present
-in an amino acid is absent or in limited supply, translation will stop at the codon specifying that amino acid
tRNA in translation
- at least one specific tRNA for each of the amino acids
- amino acids that are specified by multiple codons often have multiple tRNAs
What is the tRNA-amino acids attachment site?
CCA-3’ terminus
-if amino acid is attached, the tRNA is charged
Anticodon
Specific 3 nucleotide sequence that base pairs with the mRNA
What is the anticodon that would bind AUG?
5’-UAC-3’
Or
3’-CAU-5’
Aminoacyl-tRNA synthetases
Enzymes that attach amino acids to the corresponding tRNA
What attaches amino acids to their corresponding tRNAs?
Aminoacyl-tRNA synthetases
What does each aminoacyl-tRNA synthetase recognize?
The amino acid and ALL of the tRNAs that correspond to that amino acid
How many different aminoacyl-tRNA synthetases in humans?
20
tRNA charging
Two step reaction in which the amino acid is covalently linked via its carboxyl group to the hydroxyl group on the 3’ terminus of the tRNA (ester bond) using energy from ATP
-pyrophosphate generated and its subsequently cleaved to two molecules of inorganic phosphate
What is the energy for the amino acid attachment to tRNA?
Cleavage of two high energy phosphate bonds
Where does the amino acid attach to the tRNA?
CCA-3’ terminus of tRNA
mRNA in translation
Required as a template
Ribosomes
- Complexes of rRNAs and many proteins
- molecular machines that translate the message on an mRNA molecule into a specific protein
Eukaryotes ribosome size
80s ribosome
-60s and 40s large and small subunits
Prokaryote ribosome size
70s ribosomes
-50s and 30s large and small subunits
Ribosome subunits
Exist separately until proteins synthesis is about it being
What are the 3 ribosome sites
A, P, E
A site of ribosome
Binds incoming aminoacyl-tRNA
P site of ribosomes
Binds peptides-tRNA (peptides-tRNA carries the chain of amino acids that have already been synthesized
E site of ribosomes
- only in prokaryotes
- exit site, contains empty tRNA as it is about to exit the ribosome
Where are ribosomes located in eukaryotes?
Cytosol or bound to ER
proteins translated on the ER are destined for post-translational modifications and/or subcellular compartmentalizations and are commonly referred to as
Secretory proteins
Any protein made on the ribosome on the ER is
A secretory protein
Protein factors
Accessory proteins involved in stages of peptide synthesis
- invitation factors
- elegonation factors
- termination (release) factors
Energy requirement total for translation
Total of 4 high energy bonds are required for each amino acid that is added
Energy requirement for charging the tRNA
Two high energy bonds from ATP
Energy requirement for binding the aminoacyl-tRNA to the A site
One GTP
Energy requirement for the translocation step of translation, movement of the ribosome to the next codon
One GTP
What kind of binding between the codon and anticodon?
Antiparallel
How is mRNA read?
5’ to 3’
What would the orientation of the complementary anticodon to the mRNA be?
mRNA is read as 5’ to 3’ and the binding is antiparallel so the anticodon would be in the opposite orientation
What is the anticodon for AUG?
5’-AUG-3’
3’-UAC-5’ (rewrite it not)
5’-CAU-3’ or simply CAU
Wobble hypothesis
.tRNAs can recognize more than one codon for a specific amino acid
- the 3rd nucleotide of a codon (5’ to 3’) and the 1st nucleotide of an anticodon (3’ to 5’) can sometimes pair non-specifically
- this allows a single tRNA to recognize more than one codon
What is the net result of the wobble hypothesis?
61 different tRNAs are not absolutely required
Genetic code in wobble hypothesis
Often amino acids with multiple codons differ in the 3’ nucleotide (of the codon)
Polycistronic
Prokaryotes sometimes have multiple coding regions on the same gene
Can also have polyribosomes
Polyribosome
How many ribosomes are associated with one mRNA
Monocistronic
Euk are always this, but can have polyribosomes, slower though
When can translation being in prokaryotes?
It can being before transcription is complete
When can eukaryotic translation being?
Transcription and translation are spatially and temporally separated
How is initiation of translation potentiated in prokaryotes?
By the presence of the shine dalgarno sequence
-the 16S rRNA in the small ribose all subunit contains a sequence complementary to the Shine-Delgarno sequence- this allows correct alignment of the small ribosomal subunit with the AUG start codon
Initiation in eukaryotes (translation)
Beings with the small ribosomal subunit recognizing the 5’-cap structure and scanning along the mRNA until the first AUG is found
What makes euk monocistronic?
The mechanism of the small ribosomal subunit to recognize the 5’ cap structure and then the scanning along the mRNA until the first AUG is found
Why can prokaryotes be polycistronic?
Because the initiation process can being in the interior of the mRNA (as long as a shine-delgarno sequence and an AUG starry codon present)
Initiator tRNA
Binds the small ribosomal subunit
Initiator tRNA in prokaryotes
Bound to a formylated- methionine
Initiator tRNA in euk
Bound to a methionine (not formylated)
What is the difference in the methionine of a start codon and methionine in the polypeptide?
The start codon methionine is formylated (prok), rest are not
Elongation (translation)
The polypeptide chain is elongated by the addition of amino acids to the carboxyl end of the growing chain-forming peptide bonds
Elongation: delivery of next aminoacyl-tRNA
To the A site on the ribosome
Requires elongation factors and GTP
What is the P site on the ribosome?
The holding site
How is the formation of the peptide bond done?
By peptidyltrasnferase (a component of the large ribosomal subunit)
What plays a role in the formation of the peptide bond?
Peptidyltransferase
Where does the energy for the formation of the peptide bond come from?
Charged tRNA
Translocation
- ribosome moves three nucleotides to the next codon to be translated
- uncharged tRNA into E site
- peptiyl-tRNA into P site
- opens up A site for next aminoacyl-tRNA
- requires elongation factors and GTP
What does translocation require?
Elongation factors and GTP
Termination
When a stop codon appears in the A site
Requires GTP
Polysome/polyribsome
The mRNAs are normally much longer than the space required for a ribosome to bind, so multiple ribosomes are usually found on a single mRNA molecule
-do not confuse with polycistronic!
Secretory proteins
- model for translation
- the signal peptide is part of the polypeptide being translated
- translated at ER
- stop receptor on ER
Protein trafficking and post translational modifications
- the newly synthesized peptide emerges in the lumen of the ER and the signal peptide is cleaved
- includes secreted proteins (for exampl insulin and collagen), protein inserted into cell membranes, and others such as lysosomal enzymes
