This document summarizes key aspects of prokaryotic translation including: - Ribosomes assist in mRNA codon-tRNA anticodon binding and are composed of small and large subunits. - Translation initiation involves assembly of initiation complexes on the mRNA including ribosomal subunits and initiation factors. - Elongation adds amino acids to the growing polypeptide chain through tRNA binding sites on the ribosome and peptide bond formation. - Termination occurs when a stop codon enters the A site and release factors trigger protein release from the ribosome.

1. PROKARYOTIC
TRANSLATION
Submitted by,
Jeeva Raj Joseph
1st M.Sc. Microbiology
MSRCASC

2. Introduction TO Translation
• DNA translation is the process that converts an mRNA sequence into a
string of amino acids that form a protein.
• This fundamental process is responsible for creating the proteins that
make up most cells.
• It also marks the final step in the journey from DNA sequence to a
functional protein; the last piece of the central dogma to molecular
biology.
• The Central Dogma:

3. Ribosomes
• A structure composed of proteins and RNA (rRNA) that
assists in the binding process between mRNA codons and
tRNA anticodons.
• While ribosome can only work on one mRNA strand at a
time, multiple ribosomes can bind to any mRNA strand to
form polyribosomes.
• Eukaryotic and prokaryotic ribosomes differ slightly in
their size and complexity, though their function is
generally similar.

4. Ribosome Structure
• Ribosomes are composed of two subunits, one small and one
large.
• Four binding sites are located on the ribosome, one for mRNA
and three for tRNA. The three tRNA sites are labeled P, A, and E.
• The P site, called the peptidyl site, binds to the tRNA holding the
growing polypeptide chain of amino acids.
• The A site (acceptor site), binds to the aminoacyl tRNA, which
holds the new amino acid to be added to the polypeptide chain.
• The E site (exit site), serves as a threshold, the final transitory
step before a tRNA now bereft of its amino acid is let go by the
ribosome.

8. • The general rule of translation initiation: protein synthesis always starts
with methionine codon (AUG).
INITIATION
• Initiation of translation in prokaryotes involves the assembly of the
components of the translation system, which are: the two ribosomal
subunits (50S and 30S subunits), The mature mRNA to be translated; the
tRNA charged with N-formylmethionine (the first amino acid in the
nascent peptide), guanosine triphosphate (GTP) as a source of energy;
• The three prokaryotic initiation factors IF1, IF2, and IF3 help the
assembly of the initiation complex. Variations in the mechanism can be
anticipated.

9. • The ribosome has three active sites: the A site, the P site, and the E site.
• The A site is the point of entry for the aminoacyl tRNA (except for the
first aminoacyl tRNA, which enters at the P site).
• The P site is where the peptidyl tRNA is formed in the ribosome.
• And the E site which is the exit site of the now uncharged tRNA after it
gives its amino acid to the growing peptide chain.
INITIATION OF TRANSLATION

10. • The selection of an initiation site (usually an AUG codon) depends on the
interaction between the 30S subunit and the mRNA template.
• The 30S subunit binds to the mRNA template at a purine-rich region
(the Shine-Dalgarno sequence) upstream of the AUG initiation codon.
• The Shine-Dalgarno sequence is complementary to a pyrimidine rich
region on the 16S rRNA component of the 30S subunit.
• During the formation of the initiation complex, these complementary
nucleotide sequences pair to form a double stranded RNA structure that
binds the mRNA to the ribosome in such a way that the initiation codon
is placed at the P site.
INITIATION OF TRANSLATION

16. .
• Elongation of the polypeptide chain involves addition of amino
acids to the carboxyl end of the growing chain.
• The growing protein exits the ribosome through the polypeptide exit
tunnel in the large subunit.
• Elongation starts when the fMet-tRNA enters the P site, causing
a conformational change which opens the A site for the new
aminoacyl-tRNA to bind.
• This binding is facilitated by elongation factor-Tu (EF-Tu), a
small GTPase.
• For fast and accurate recognition of the appropriate tRNA, the
ribosome utilizes large conformational changes (conformational
proofreading) .
ELONGATION

17. ELONGATION IN PROKARYOTES
• Now, the A site has the newly formed peptide, while the P site has an
uncharged tRNA (tRNA with no amino acids).
• Now, with tRNA bearing a chain of amino acids in the p site and tRNA
containing a single amino acid in the A site, the addition of a link to the
chain can be made.
• This addition occurs through the formation of a peptide bond, the
nitrogen-carbon bond that forms between amino acid subunits to form a
polypeptide chain.
• This bond is catalyzed by the enzyme peptidyl transferase.

19. ELONGATION IN PROKARYOTES
• The newly formed peptide in the A site tRNA is known as dipeptide
and the whole assembly is called dipeptidyl-tRNA.
• The tRNA in the P site minus the amino acid is known to be
deacylated.
• In the final stage of elongation, called translocation, the deacylated
tRNA (in the P site) and the dipeptidyl-tRNA (in the A site) along
with its corresponding codons move to the E and P sites,
respectively, and a new codon moves into the A site.
• This process is catalyzed by elongation factor G (EF-G).

21. ELONGATION IN PROKARYOTES
• The deacylated tRNA at the E site is released from the ribosome during
the next A-site occupation by an aminoacyl-tRNA again facilitated by EF-
Tu.
• The ribosome continues to translate the remaining codons on the mRNA
as more aminoacyl-tRNA bind to the A site, until the ribosome reaches a
stop codon on mRNA(UAA, UGA, or UAG).
• In bacteria, translation initiation occurs as soon as the 5' end of an mRNA
is synthesized, as translation and transcription are coupled. This is not
possible in eukaryotes because transcription and translation are carried out
in separate compartments of the cell (the nucleus and cytoplasm).

23. Termination
• Termination occurs when one of the three termination codons moves
into the A site.
• These codons are not recognized by any tRNAs.
• Instead, they are recognized by proteins called release factors, namely
RF1 (recognizing the UAA and UAG stop codons) or RF2
(recognizing the UAA and UGA stop codons).
• These factors trigger the hydrolysis of the ester bond in peptidyl-tRNA
and the release of the newly synthesized protein from the ribosome.
• A third release factor RF-3 catalyzes the release of RF-1 and RF-2 at
the end of the termination process.

27. • Proteins in prokaryotes are synthesized at a rate of only 18 amino acid
residues per second, whereas bacterial replisomes synthesize DNA at a
rate of 1000 nucleotides per second.
• This difference in rate reflects, in part, the difference between
polymerizing four types of nucleotides to make nucleic acids and
polymerizing 20 types of amino acids to make proteins.
• Testing and rejecting incorrect aminoacyl-tRNA molecules takes time
and slows protein synthesis.
Translation machinery works relatively slowly compared to the
enzyme systems that catalyze DNA replication

28. Submitted to,
Dr.VemulaVani
Department Of Microbiology
MSRCASC
Thank you