Transcription• RNA Polymerase ‘reads’ template DNA strand to make complement RNA (mRNA code for a polypeptide).• Associated with a gene(s) is an up-gene promoter/operator sequence for RNA polymerase binding and a down-gene termination sequence.• Gene transcription can be regulated (on/off switch) negatively or positively by regulatory proteins (more later).

Translation• Ribosome moves along the mRNA, codon-by-codon, adding appropriate amino acids via specific tRNAs to its growing polypeptide product.

Transcription

Trailer sequence

Operator region

Gene(s)

Sigma-factor

Genes + p/operator = operon.

Antisense (template) strand

Sense (complementary) strand

3’

5’

5’ antisense

3’ sense

Promoter Region

Pribnow Box

• Site for RNA Polymerase recognition and binding.• Recognition requires involvement of a sigma factor.• Different promoter types have their own sigma factor.• There can be an adjacent regulator binding site (= operator).

mRNA Elongation(recall that Us replace Ts)

5’

3’

5’

3’

Transcription Termination

• Rho – dependent: a protein called “ρ” unwinds the DNA template – mRNA complex; thus weakening the association until mRNA and RNA polymerase falls off DNA.

• Rho-Independent: here two G+C regions rich slow RNA polymerase; a hairpin forms in the mRNA weakening the mRNA-DNA template association; which is further weakened by poly-A stretch.

rRNA and tRNA Products• rrn-gene has multiple rRNA & tRNA products.• Spacer RNA is spliced out by special nucleases.• rrn-operons are often located close to Ori.• Why might this have been selected? (think fast growth)?

mRNA product• Within the leader is the Shine-Dalgarno sequence (5’AGGA3’). It complements a site on 16SrRNA of ribosome; used to bind a ribosome to mRNA for translation.

• Trailer at 3’ end of mRNA was required for proper ending of transcription.

• Translation of mRNA information into protein starts at the initiation codon (5’AUG3’); proceeds codon by codon until the end of gene, as identified by a stop, or non-sense, codon.

• Coding region of mRNA can be polygenic (multiple genes = multiple starts / stops).

Direction of Translation

Coding Region

Genetic Code: (3 nonsense; 61 coding yet fewer tRNA)

start

Genetic code degeneracy; more than one codon for some amino acids.

Results from tRNA “wobble” of first anticodon position; sometimes a novel nucleotide like inosine (e.g. ICC anticodon for Gly codons GGA, GGC, GGU).

3’-A U G-5’Codon written in reverse (3’→5’)

Initiation Codon establishes the proper “reading-frame”!

Translation

Translation Initiation

Three initiation factor proteins are involved (IF1, IF2, IF3).

30S subunit alone recognizes Shine-Delgarno sequence of mRNA, which binds initiation tRNA, IF1 & IF2; IF3 released.

50S subunit binds 30S (=70S), sandwiching mRNA and placing initiation tRNA at the P-site.

A-site ready to receive tRNA for next codon. Ribosomes slide along mRNA in 5’→3’ direction.

Translation ElongationPeptidyl

transferase

Translation Termination

Stop codon

Elongation of polypeptide continues until a stop codon is reached at the A-site.

Ribosome stalls, allowing release factor proteins to bind.

Upon dissociation of 50S from 30S, the IF3 binds 30S to make it receptive to a new mRNA start.