Transcription: RNA synthesis and Processing Flashcards
Different types of RNA
Messenger RNA: encodes the AA sequence of all polypeptides found in the cell (5% of total rna) most complex
Transfer RNA: matches specific AA to triplet codons in mRNA during protein synthesis (15% of total RNA)
Ribosomal RNA: constituents and catalytic appropriate AA, 80% of total RNA not complex
Ribonucleic acids play several roles other than information
microRNA: non coding involved in regulating the expression of genes
RNA metabolism
RNAs are synthesized in cells using DNA as the template in transcription. Transcription in tightly regulated in order to control the concentration of each proteins
Many RNAs can fold into compact structures with specific functions (mainly due to the fact that its mostly ssRNA). Some RNA molecules can act as catalysts (ribozymes), often using metal ions as cofactors
Most eukaryotic RNAs are processed after synthesis (eliminations of introns and joining of exons, poly A at the 3’ end and capping the 5’ end)
The basic reaction of Transcription
Some nucleoside is added to the free 3’ Oh bc even RNA is synthesized in the 5’-> 3’ direction
3’OH of the existing RNA chain undergoes a nucleophilic attack on the alpha phosphate of the incoming nucleotide
Growing chain is complementary to the template strand
Synthesisi is catalyzed by RNA polymerase
Properties of the RNA polymerase
In prokaryotes a single RNA polymerase polymerizes mRNA, rRNA and tRNA
Prokaryotic RNA polymerase is a multisubunit enzyme
RNA polymerase has 5 core subunits and a sigma subunit
Core polymerizes, Sigma factor allows the recognition of promoter regions on the DNA
RNA polymerase lacks the 3’->5’ exonuclease activity so it has a high error rate but its also very fast
RNA binds to promoter regions to initiate transcription
Bacterial gene
Promoter: site for binding RNAP
Operator: binding sites for repressor or activator proteins
Structural genes: sometimes many genes per single unit
The whole thing is called an operon
RNA initiation, elongation and termination
- RNAP core binds to the DNA promoter (Promoter, Polymerase)-sigma binds to the TATA box
- Transcription bubble forms (closed complex-wound DNA)
- Transcription is initiated (open complex-unwound DNA after RNAP binds and unwinds DNA)
- Promoter clearance
- Elongation
Terminology of Transcription
Template Strand- the strand that the RNA is attached to
Non template/ Coding Strand- the strand that the RNA is no attached to but its the exact copy of it because its copying the sister strand
Transcriptional elongation
Occurs in the 5’ to 3’ direction using the energy of the phosphate bond from the incoming Ribonucleotides, the sigma factor dissociates from the holoenzyme after elongation initiates.
Multiple RNA plymerase complexes load onto promotor region in sequential fashion, very proccessive
positive super hilicity occurs infront of the transcription bubble and releived via topoisomerases
Specificity of transcription
different sigma factors recognize different promoter regions
sigma increases specificity of RNAP but decreases affinity
you can change the sigma via sigma factors to lead to diferent gene expression, change in sigma utilization can be used to regulated
strong promoters cause frequent initiation and tend to conform closely to the consenses
Termination of Transcription
when theres a hairpin loop formed by the palindrome sequence (GC), theres a bunch of As from DNA and a bunch of Us from RNA that are bound and that A-U bond is weaker than the bond that would be formed from the A-T (the Ts are in the coding/non template strand) and so the DNA comes back together, popping off the RNA transcript
occurs by protein independent or protein dependent mechanisms both rely on the hairpin loop and AU dissociation
Protein dependent termination of transcription
Terminator protein Rho interacts with RNAp when hairmin starts
Rho is an ATP dependent RNA/DNA helicase-> dissociation hybrid
Rho binds to RUT site
3 different kinds of eukaryotic RNA polymerases
Each one synthesizes a different class of RNA
RNA polymerase 1 synthesizes rRNA
RNA polymerase 2 synthesizes mRNA (super fase, inhbitied by alpha amantin) can recognize thousands of promoters
RNA polymerase 3 makes tRNAs and small RNA products
Mitochondria have their own polymerase
Eukaryotic Post transcriptional processing of precursor mRNA in Eukaryotes
in the gene there are coding regions-> the actual gene and there are noncoding regions called intervening sequences
Theres a 3’ UTR region thats not for coding
1st you transcribe a primary transcript with POL 2 then the primary transcript gets processed into mature mRNA
The processing includes:
5’ cap
3’ poly a tail on the UTR
splicing out introns (non coding regions)
The addition of the 5’ Cap
Guanylyl transferase adds a 7 methyl guanine on the end (Using SAM as the donor)
- enhances stability-protection from nucleases
- enhances translation efficiencey
The addition of the poly A tail
Important for mRNA stability
Helps in translation
Added downstream of the poly A signal in 3’UTR
it adds a lot of As
