Transcription and Translation: Initiation & Termination

Depends on the recognition of mRNA by ribosomes.

• This recognition begins at a specific sequence, the Shine-Dalgarno Sequence (5’ AGGAGG 3’). The first AUG after the Shine-Dalgarno interacts with fMETHIONINE-tRNA and initiates translation.

Because of multiple Shine-Dalgarno sequences, bacterial mRNA can be polycistronic (coding for more than one protein) while eukaryal mRNA is monocistronic.

The Shine-Dalgarno sequence attaches to the mRNA through base-pair interactions between the 16s rRNA and the SD sequence.

• Mutations within the Shine-Dalgarno sequence can eliminate or greatly reduce the translation of mRNA.

• Mutations within the 16s rRNA can affect overall protein synthesis within the cell.

• A specific mutation within the SD can be corrected by a complementary mutation in the 16S rRNA sequence.

Following the initiation, the ribosome moves along the mRNA reading the codons. With each successive codon, a tRNA with a complementary anticodon (charged appropriately) is recruited and enters the aminoacyl site.

• Peptide bonds form between adjacent amino acids resulting in polypeptide formation.

• Once the ribosome encounters a stop codon, translation ceases and the polypeptide leaves the ribosome.

Mostly the same as bacteria except for 3 differences:

• A non-modified methionine is used and initiator tRNA recognizes the start codon.

• Eukaryal mRNAs do not contain a Shine-Dalgarno-like sequence. It just initiates translation at the first AUG on the 5’ end.

• They also have many more initiation factors. A cap-binding protein (CBP) first binds to the 5’ cap and then additional polypeptides bind before the ribosome can attach.

Small, 40S subunit containing:

• 33 polypeptides

• 18S rRNA

Small, 30S subunit containing:

• 21 polypeptides

• 16S rRNA

Large (60S) subunit

• 49 polypeptides

• 28S rRNA

• 5.8S rRNA

• 5S rRNA

Large (50S) subunit

• 31 polypeptides

• 23S rRNA

• 5S rRNA

The core enzyme is a complex of β, β′, ω, and two copies of σ.

• The core enzyme interacts with the Sigma factor to initiate transcription by forming holoenzyme.

• This enzyme interacts with the promoter where RNA polymerase binds and helps unwind the DNA and creates a molecule of RNA.

• Transcription does not begin at the promoter.

The site of transcription initiation is referred to as the +1 position and, in bacteria, typically is an A or a G.

A TATA box is a DNA sequence that indicates where a genetic sequence can be read and decoded. It is a type of promoter sequence, which specifies to other molecules where transcription begins. Transcription is a process that produces an RNA molecule from a DNA sequence.

RNA polymerase II does not bind directly to DNA. Rather, its binding is facilitated by various transcription factors, including TATA-binding protein (TBP) and transcription factor IIB (TFIIB).

The promoters in eukaryal structural genes consist of several motifs, including TATA, GC, CAAT, and octamer (eight nucleotide) boxes.

Transcription factors TBP and TFB interact with the promoter.

• RNA polymerase is recruited.

• Unwinding of the DNA occurs and transcription begins.

Rho-dependent: Rho factor binds to the RNA molecule and travels along the molecule. The RNA polymerase will slow down and pause when it reaches a specific termination site, and the Rho factor replaces the RNA polymerase causing transcription to end.

Rho-independent: Intrinsic termination ends with a short segment of GC-rich repeats, followed by a string of A’s.

• The hydrogen bonds in the RNA molecule allow a hairpin structure to form.

• This hairpin structure causes the RNA polymerase to fall off the DNA, and the A:U are so weakly bonded to each other that they that the RNA molecule dissociates.

Termination of transcription by RNA pol I resembles the rho-dependent mechanism.

Termination of transcription by RNA pol III resembles rho-independent mechanism.

While for RNA pol II undergoes three major forms of processing:

• A 7-methylguanosine cap is added to the 5’ end via a 5’ to 5’ triphosphate linkage.

• A poly A tail is added to the 3’ end.

• Introns are spliced out, bringing together the exons.