Bacterial Transcription and translation

What is transcription?

information in DNA strand copied into mRNA

What is translation?

mRNA is decoded by a ribosome to produce a specific polypeptide or protein.

Describe the bacterial structural of a gene that help initiate transcription

• regulatory structures/sequences upstream of the coding region that control expression. include promoter sequence

• sigma factor and RNA polymerase recognition sites enable the specific binding of RNA polymerase

• The Pribnow box (-10 box) - conserved TATAAT sequence, located 10 base pairs upstream from the transcription start site is essential for RNA polymerase binding and initiation of transcription. 

describe the flow from DNA to mRNA

• dsDNA (double stranded)

• transcription

• mRNA

• always 5’ to 3’

Descfribe the 4 stages of transcription

1. Pre-initiation - sigma factor directs RNA polymerase core enzyme to -35

2. Initation - RNA polymeraze denatures/separates DNA at -10 site, stabilized by sigma factor (protein subunit of RNA polymerase)

3. Elongation - RNA polymerase synthesizes RNA, sigma factor detaches

4. Termination - RNA polymerase encounters terminator sequence

What is the UTR?

untranslated region in mRNA

What is the promoter region?

region of DNA recognized by sigma factor, initiates transcription

What is the promoter region and how does it work?

• region of DNA recognized by sigma factor, initiates transcription

• ATG pattern to start

• sigma recognizes -35 portion of DNA, helps RNA polymerase bind to template

• sigma and RNA polymerase “open” dsDNA at -10 with helicase

What is intrinsic termination?

• mechanism in bacterial transcription where RNA polymerase detects A-rich terminator sequence in the newly synthesized RNA that signal it to stop transcription

• cause formation of a hairpin loop

• destabilizes bonds and releases RNA polymerases

Describe the initaiton of transcription

• region of DNA recognized by sigma factor, initiates transcription

• sigma recognizes -35 portion of DNA, RNA polymerase binds to template

• sigma and RNA polymerase “open” dsDNA at -10

• sigma released during transcription

• different sigma factors with different binding sequences bind different promoters

What are the mechanisms for termination of transcription?

• intrinsic termination - step loop secondary structure

• rho factor dependent terminatino

What is rho factor dependent termination?

• termination of transcription mechanism

• rho protein binds to rho recognition site in mRNA and chases RNA polymerase

• RNA polymerase pauses

• rho catches up, separates RNA-DNA hybrid using helicase

What is translation?

The process of synthesizing proteins from mRNA sequences, where ribosomes read the mRNA codons and tRNA molecules bring the corresponding amino acids.

Describe the structure of tRNA

• amino acid attaches at 3’ end

• antiocodon at bottom, contains 3 nucleotides complementary to mRNA codon, allowing tRNA to read mRNA and deliver correct polypeptide

What are the steps of translation initiation?

• Fmet amino acid on end of first tRNA

• Initiation factor IF3 prevents small ribosome subunit (30s) from binding to large subunit (50s)

• IF2 binds GTP and fmet-tRNA, guides them to P site on 30s subunit

• IF1 binds 30s, kicking off IF3, kicking off !F3 initiation complex so it can bind 50s

• GTP bound to IF2 hydrolyzed, all IFs released

• 70s complex formed, fMet TRNA in P site and A site open

What are the steps of the translation elongation cycle?

• Binding - EF-Tu binds GTP and tNA, brings to A site

• Transpeptidation - peptide bond forms between p site and a site amino acids

• Translocation - peptide chain on A site, tRNA noves to p site, other leaves out the E site

What are the p site, A site, and e site?

• sites in ribosome for translation

• A site - arrival - entrypoint for incoming tRNA carrying an amino acid

• p site - peptide bond formed between new amino acid and chain; first tRNA binds here during initiation of translation

• E site - exit - empty tRNA exits

What is transpeptidation?

transfer of growing polypeptide from P site to A site, peptide bond formed

How is translation terminated

• release factor proteins bind to UAA/UAG/UGA stop codons

• polypeptide released

• ribosome dissociates

What is couple transcription and translation in bacteria?

• translation starts before transcription is finished

• Multiple ribosomes attach to a single mRNA strand simultaneously, called a polyribosome

• allows bacteria to respond quickly to environmental changes by rapidly producing proteins.

Describe how bacteria translocate proteins to outside

• signal peptides at the amino terminal of protein act as tag for secretion

• most bacteria use SEcA system with SEC protein pushing protein out of a membrane channel

• some use TAT system

Where does transcription of DNA start in bacteria?

ATG promoter region, called -35 region. sig

What is fMet?

modified amino acid used exclusively by bacteria to initiate protein synthesis during translation.

Why is translocation (secretion or inserting in membrane) protein to the ourside important?

can occur during host pathogens interactions

What is the leader sequence in mRNA ?

• mRNA region thats transcribed but not translated before the start codon

What are the 3 primary regions of mRNA and their functions?

• leader - transcribed but not translated

• shine-dalgarno - site within leader region where translation begins

• coding region - protein region

What is the Shine-Dalgarno sequence?

site in mRNA leader sequence where translation begins