Prokaryotic Translation and Transcription

Transcription

Transcription is the process of transcribing one strand of DNA into a complementary strand of RNA . It is catalyzed by the enzyme RNA polymerase. It takes place in the direction from the 5' end of the new RNA molecule to its 3' end.

Transcription in prokaryotes follows a similar basic mechanism to that of eukaryotes . For more information with an explanation, see the article Transcription in Prokaryotes and the Operon Model .

RNA polymerase

Transcription in bacteria is ensured by a single RNA polymerase:

• 5 protein subunits α, β, β', ω and σ; α in two copies
• The σ subunit is used to bind n DNA, it dissociates after the start of synthesis

Initiation

Initiation of RNA synthesis occurs in bacteria as follows:

• promoter recognition using the sigma factor (multiple promoter variants + σ)
• σ recognizes −35 and −10 (TATA box/Pribnow box) of the consensus sequence
• opening of the transcription bubble
• synthesis of a short chain around 9 nt, dissociation of the σ factor
• energetically, transcription is driven by hydrolysis of the macroergic bond of the incoming ribonucleoside triphosphate

Termination

Transcription termination of prokaryotes can be Rho (ρ) independent or ρ dependent:

– Rho independent termination:

• a termination sequence containing inverted repeats separated by a non-repetitive stretch rich in GC pairs
• forming a transcription-stopping loop structure
• the following stretch of polyA (in DNA) allows easy dissociation

– Rho dependent

• The Rho factor recognizes a sequence in RNA
• it moves towards the polymerase and causes dissociation

Translation

Translation is the process of synthesizing a polypeptide chain based on the information contained in mRNA . Triplets of nucleotides in RNA are translated into the form of individual amino acids of a polypeptide according to the rules of the genetic code . Translation in prokaryotes follows a similar basic mechanism to that of eukaryotes . Further information with an explanation can be found in the article Translation in prokaryotes , Translation and the question Translation in eukaryotes , the basic features and specificities of prokaryotic translation are presented here.

Aminoacyl-tRNA synthesis

The tRNA molecule brings the amino acid residue to the ribosome. Bacterial tRNAs are characterized by:

• roughly 60 types of tRNA (versus 100–110 in a mammalian cell)
• 73–93 nucleotides in length
• secondary structure of the shape of a four-leaf clover, tertiary structure of the letter L
• acceptor arm terminated by a CCA triplet
• dihydrouridine arm (D or DHU), pseudouridine arm (T or TΨC), variable arm
• anticodon arm

Activation of tRNA by binding to an amino acid residue occurs through the enzyme aminoacyl-tRNA-synthetase with the consumption of ATP:

• amino acid + ATP ↔ aminoacyl-AMP + pyrophosphate
• aminoacyl-AMP + tRNA ↔ aminoacyl-tRNA + AMP

The reaction is thermodynamically driven by pyrophosphate decomposition.

Prokaryotic ribosome

The bacterial ribosome, like the eukaryotic ribosome, consists of two subunits:

– 30S subunit

• 16S rRNA (with 3' end complementary to Shine-Delgarno sequence)
• 21 proteins

– 50S subunit

• 5S rRNA, 23S rRNA (with peptidyltransferase activity – catalyzes peptide chain elongation)
• 31 proteins

Translation initiation

Proteosynthesis itself can be divided into initiation, elongation and termination. Bacterial initiation proceeds as follows:

• the first amino acid of most bacteria is N-formylmethionine attached to the special tRNA ^fMet . Usually 1–3 N-terminal amino acids are cleaved post-translationally.
• N-formylmethionyl-tRNA ^fMet binds to the free 30S subunit of the ribosome
• The mRNA is bound by the interaction of the 3' end of the 16S rRNA with a Shine-Delgarno sequence near the 5' end (RBS - ribosome-binding site), usually 8 nucleotides from the initiation codon AUG
• fMet-tRNA interacts by anticodon with initiation codon AUG (sometimes GUG)
• The 50S subunit of the ribosome binds so that the fMet-tRNA is in the P site of the ribosome
• initiation of translation requires initiation factors IF1, IF2 and IF3 and consumes energy in the form of GTP

Elongation

During elongation, amino acid residues are added to the C-terminus of the polypeptide through a repeating sequence of events:

• The P site of the ribosome (see translation ) is occupied by N-formylmethionyl-tRNA^fMet or peptidyl tRNA, and the A site is empty
• aminoacyl-tRNA corresponding to the following codon binds to the A site with the help of EF-Tu factor and GTP consumption
• a transpeptidase reaction catalyzed by 23S rRNA takes place – the α-amino group of the amino acid residue in the A site nucleophilically attacks the α-carboxyl group of the C-terminal amino acid in the P site. The peptide thereby moves to the tRNA in the A site. The reaction does not consume any energy-rich molecules.
• translocation occurs: the peptidyl-tRNA moves from the A site to the P site, the ribosome moves one codon to the mRNA, and the previous tRNA moves to the E site. The process requires the EF-G factor and the hydrolysis of a GTP molecule
• The tRNA at the E site leaves the ribosome and the cycle repeats

Termination

Termination of translation in a prokaryotic cell occurs as follows:

• one of the three (usually) termination (nonsense) codons – UAA, UAG or UGA – gets to A instead of the ribosome
• is recognized by one of three termination factors (RF-1, RF-2 or RF-3)
• the peptide is hydrolytically released by peptidyl transferase activity
• dissociation of ribosome subunits and translation proteins occurs.
• IF-3 remains bound to the 30S subunit, preventing reassociation with the 50S subunit

Links

Related articles

• Transcription
• Transcription in Prokaryotes
• Translation in Prokaryotes
• Translation in Eukaryotes
• Translation
• Regulation of gene expresion in Prokaryotes
• RNA
  • mRNA

References

• MADIGAN, Michael – BENDER, Kelly – MARTINKO, John, et al. Brock Biology of Microorganisms. - edition. Pearson, 2014. pp. 1030. ISBN 9781292018317.

• PRESCOTT, Lansing – HARLEY, John – KLEIN, Donald. Microbiology. - edition. WCB/McGraw-Hill, 1999. pp. 963. ISBN 9780697354396.

• KOHOUTOVÁ, Milada, et al. Lékařská biologie a genetika (II. díl). 1. edition. Karolinum, 2012. ISBN 978-80-246-1873-9.