L11 - Translation

Question 1

The codon facts

Answer 1

Three bases encode an amino acid - codons
Code is non-overlapping and degenerate
- Some amino acids are specified by more than one codon (61 codons and 20 amino acids)
Three possible reading frames from one mRNA

Question 2

Start codon

Answer 2

AUG = Methionine

Question 3

Stop codons

Answer 3

UAA UAG UGA - signal end of open reading frame

Question 4

tRNA base pairing

Answer 4

One end base pairs with codon - anticodon loop
Other end carries the amino acid - 3’ end
Intermolecular base pairing within tRNA gives it structure

Question 5

Nucleotides within tRNA

Answer 5

Primary sequences of nucleotides varies, even within double stranded regions
Some nucleotides in tRNAs are modified to allow different interactions

Question 6

What is the ratio of tRNA to codon?

Answer 6

Not a 1:1 ratio
Wobble bases (position 3) allow same anticodon to bind to more than one codon
One way wobble is made is by modification

Question 7

Types of wobble modification

Answer 7

• Deamination of A to create an inosine

- Inosine can pair to U, C or A

Question 8

Method of coupling amino acid to the tRNA

Answer 8

1. Aminoacyl-tRNA synthetases primes amino acid by adding AMP to C-terminus
2. Uses the adenylated amino acid to form aminoacyl-tRNA
   - Known as charged tRNA - energy from ATP hydrolysis is contained in ester linkage

Question 9

What are the two adapters required for translation?

Answer 9

Synthetase

tRNA

Question 10

Synthetase adapter

Answer 10

Pairs correct amino acid to correct tRNA
They are specific to individual tRNAs
Amino acids have to fit into two pockets in the synthetase - before and after AMP addition
Nucleotides in the anticodon and acceptor stem have pockets in the synthetase

Question 11

tRNA adapter

Answer 11

Pairs correct codon to correct amino acid within ribosome

Pairing requires specific interactions between molecular surfaces

Question 12

Ribosome subunits

Answer 12

Two subunits

• Large subunit - catalyzes polymerization
• Small subunit - facilitates tRNA/mRNA interaction

Question 13

Method of ribosome function

Answer 13

1. Subunits come together on 5’ end of mRNA
2. Process along mRNA at two amino acids per second
3. Separate at the stop codon

Question 14

How are new amino acids added to the ribosome?

Answer 14

New amino acids are added to the C-terminus of the protein by peptidyl transferase
Replace high energy bond with low energy bond

Question 15

Method of movement through ribosome

Answer 15

1. Charged tRNAs enters A-site
2. Petidyl transferase catalyses amino acid addition
3. Conformational changes move tRNAs to E- and P-sites and move small subunit three nucleotides
4. Uncharged tRNAs leave E-site

Question 16

How many tRNAs are in the ribosome at once?

Answer 16

2

Question 17

What is the role of elongation factors?

Answer 17

Help translation and improve accuracy

Question 18

EF-1 role

Answer 18

Once the anticodon is bound, EF-1 causes two delays before petidyl transferase can act
Hydrolyses GTP to GDP
- More rapid if the codon and anticodon are correctly matched
Dissociates from tRNA
These lags allow time for incorrectly bound tRNAs to fall off
- Some correct tRNAs also fall but at a slower rate

Question 19

Ribosome structure

Answer 19

Large subunit rRNAs form a structure that contains most of the catalytic activity
- Including petidyl transferase
Riboproteins lie on the surface

Question 20

Which amino acid tRNA assembles the ribosome

Answer 20

AUG – methionine

Question 21

Method of ribosome assembly

Answer 21

mRNA that has a cap and tail is bound by eIF-4G and eIF-4E to form a loop
- Checkpoint for broken mRNA
- Eukaryote Initiation Factors
Only the methionine tRNA with eIF-2 can bind to small ribosome subunit alone
- Complex binds to cap and associated initiation factors
- Scans along mRNA and settles on the first AUG
- eIF2 is released and ribosome forms

Question 22

What are stop codons recognised by?

Answer 22

Recognized by release factors

• Look like charged tRNAs and enter the A-site
• Results in dissociation of the ribosome

Question 23

How far are ribosomes spaced apart on the polysome?

Answer 23

80 nucleotides

Question 24

How does protein folding occur?

Answer 24

Folds rapidly putting hydrophobic side chains in the middle
- Achieves a lower energy state
Multistep process - important that steps occur in right order
- Incorrect step may reduce the energy state but block further folding

Question 25

Misfolded proteins

Answer 25

Generally have exposed hydrophobic regions - lead to aggregation

Question 26

What is it called when proteins initially fold into roughly the correct confirmation?

Answer 26

Molten globule

Question 27

What is the role of molecular chaperones?

Answer 27

Reverse incorrect steps in protein folding

Question 28

What are the two types of heat shock protein?

Answer 28

hsp60 – put misfolded proteins into isolation | hsp70 - bind to exposed hydrophobic amino acids on proteins

Question 29

How do heat shock proteins work?

Answer 29

High temperatures cause properly folded proteins to unfold | - Chaperones function during normal folding as well as when cell has been overheated

Question 30

HSP60 family

Answer 30

1. Hydrophobic entrance binds to protein partially unfolding it 2. GroES cap seals protein inside for 15 seconds to allow refolding Polyubiquitination marks for destruction in the proteasome 1/3 of all newly synthesized proteins are immediately recycled

Question 31

Protein aggregates can cause?

Answer 31

Disease They are large and protease resistant - Can lead to cell death - Can cause a chain reaction to misfold more proteins

Question 32

CJD, Huntington’s and Alzheimer’s

Answer 32

Associated with large, extracellular protein aggregates

Question 33

Amyloid plaques

Answer 33

Made up of cross-beta filaments

Question 34

CJD prions

Answer 34

Convert normal proteins | Animals eat infected tissue and some prions enter brain and seed new cross-beta filaments