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Proteins translated on ribosomes associated with the rough endoplasmic reticulum:
Secreted proteins
Proteins inserted into the cell membrane
Lysosomal enzymes

Proteins translated on free cytoplasmic ribosomes:
Cytoplasmic proteins
Mitochondrial proteins

Translation

1

Each codons consists of 3 bases (triplet)
There are 64 codons
They are all written in the 5' to 3' directions

61 codons code for amino acids
The other 3 (UAA, UGA, UAG) are stop codons (nonsense codons) that terminate translation

There is one start codon (initiation codon), AUG coding for methionine
Protein synthesis begins with methionine in eukaryotes and formylmethionine in prokaryotes

The Genetic Code

2

Characteristics of the Genetic Code

Specific

Universal

Redundant/Degenerate

Nonoveerlapping/Commaless

3

A specific codon always codes from the same amino acid

Specific

4

It has been conversed from very early stages of evolution with only slight differences in the manner in which the code translated

Universal

5

A given amino acid may have more than one triplet coding for it

Redundant/Degenerate

6

Code is read from a fixed starting point as a continuous sequence of bases, taken 3 at a time

Nonoverlapping/Commaless

7

Requirement of Translation

1. All the amino acids that eventually appear in the protein

2. At least one specific type of RNA for each amino acid

3. One aminoacyl-tRNA synthetase for each amino acid

4. mRNA coding for the protein to be synthesized

5. Fully competent ribosomes

6. Protein factors

7. ATP and GTP as energy sources

8

Has an attachment site for a specific amino acid at its 3'end

Also has an anticodon region that can recognize the codon specifying the amino acid the tRNA is carrying

tRNA

9

Accurate base pairing is required only in the first 2 nucleoside positions of an mRNA codon, so codon differing in the 3rd wobble position may code for the same tRNA/amino acid

tRNA Wobble

10

Large complexes of protein and rRNA

Consists of 2 subunits

Eukaryotes 60s and 40s = 80s

Prokaryotes 50s and 30s = 70s

Ribosomes

11

Each ribosomes has 3 binding sites

A site codon
P site codon
E site codon

12

Binds an incoming aminoacyl-tRNA

A site codon

13

Occupied by peptidyl-tRNA

P site codon

14

Occupied by the empty tRNA as it is about to exit the ribosomes

E site codon

15

Amino-acetyl-tRNA synthetase (1 per AA) uses an ATP scrutinizes an AA before and after it binds to tRNA

If incorrect, bond is hydrolyzed by synthetase. The AA-tRNA bond has energy for formation of peptide bond

A mischarged tRNA reads usual codon, but inserts wrong AA

Charging

16

Steps in Translation

Step 1 : initiation
Step 2 : elongation
Step 3 : termination

17

Activated by GTP hydrolysis, initiation factors (elFs) help assemble the 40s ribosomal subunit with the initiator tRNA and are released when the mRNA and the ribosomal unit assemble with the complex

In prokaryotes, a purine-rich region (the SHINE-DALGARNO SEQUENCE) of the mRNA base pairs with a complementary sequence

The 5'-cap in eukaryotic mRNA is used to position that structure on the ribosome

INITIATION

18

1. Aminoacyl-tRNA binds to A site (except for initiator methionine). Elongation factors direct the binding of the appropriate tRNA to the codon in the empty A site

2. The enzyme peptidyltransferase catalyzed peptide bond formation, transfers growing polypeptide to amino acid in A site

3. Ribosome advance 3 nucleosides toward 3' end of RNA, moving peptidyl tRNA to P site (translocation)

ELONGATION

19

Releasing factors are proteins that hydrolyze the peptidyl-tRNA bond when a stop codon (UAA, UAG, UGA) occupies the A site

Completed protein is released from ribosome through simple hydrolysis and dissociates

TERMINATION

20

Energy requirement of Translation

1. tRNA aminoacylation (ATP -->AMP)
2. Loading tRNA onto ribosome (GTP --> GDP)
3. Translocation (GTP --> GDP)

21

Post-translational modification

Trimming excess amino acids
Phosphorylation
Glycosylation
Hydroxylation
Proteins that are defective or destined for rapid turnover are marked for destruction by ubiquitin and are degraded by proteasomes

22

An exotoxin of CORYNEBACTERIUM DIPHTHERIAE inactivates eEF-2 and thereby specifically inhibits mammalian protein synthesis

Diphtheria toxin