1. Medical Genetics-Transcription and Translation Robert F. Waters, PhD
DNA
hRNA
mRNA
rRNA
tRNA

2. DNA
Helical Structure
Base Pairing
Chromatin/Chromosomes
Operon

3. Protein Synthesis-Overview
Major classes of RNA
mRNA
Prokaryotic and Eukaryotic
Eukaryotic (hnRNA – mRNA)
Leader (Header) region
5’ untranslated region (5’ UTR)
Eukaryotic cap region attached 7-methylguanylate
Trailer region
3’ untranslated region (3’ UTR)
Amount of mRNA
About 5% in prokaryotes
Stability
mRNA lasts for just a few minutes

4. RNA Ribosomal RNA (rRNA) Prokaryotic Three kinds
23s rRNA (2904 nucleotides)—component of 50s rRNA subunit
16s rRNA (1541 nucleotides)—part of small 30s rRNA subunit
5s rRNA (120 nucleotides)—part of 50s rRNA subunit
Abundance – about 80% of all RNA

5. RNA Continued: Ribosomal RNA Eukaryotic Abundance 28s (60s subunit)
Transcription product of separate gene
Abundance
4% -- 45s
71% -- full rRNA

6. RNA Continued: tRNA (Transfer RNA) Prokaryotic
Average about 80 nucleotides
Some common structure so may function with rRNA
tRNAs derived from larger precursor tRNAs
Abundance (about 15% of total RNA in prokaryotes)

7. RNA Continued: tRNA (Transfer RNA)
Eukaryotic (Very similar to prokaryotic)
Average about 80 nucleotides
Some common structure so may function with rRNA
tRNAs derived from larger precursor tRNAs
Abundance (about 15% of total RNA in prokaryotes)

8. Other RNAs Small RNAs Eukaryotes Small Cytoplasmic RNAs (scRNAs)
Signal recognition particles
Small Nuclear RNAs (snRNAs)
Snurps
Spliceosome
Separation of introns and exons

9. Transcription Formation of RNA from DNA Nucleus
Single strand of DNA acts as template
Substrates
ATP
GTP
CTP
UTP
Note cancer chemotherapy

10. Direction of RNA Synthesis
RNA chain grown proceeds from 5’ to 3’
Codon sequence
Therefore amino acid sequence
Not understood as to which DNA strand

11. Elongation Enzymes in Transcription
Prokaryotes
DNA Dependent RNA Polymerase
Core enzyme is trimeric
Necessary for RNA elongation
Holoenzyme has core enzyme with another subunit required for initiation
Eukaryotes
Many large enzymes usually greater than MW
Called Type I, II, III

12. Promoters of Transcription
Prokaryotic Promoter sequences
Initiation site (startpoint)
Pribnow Box (similar to TATA box)
TATAAT
9-18 BP upstream from initiator
-35 sequence
15 BP upstream from Pribnow Box
TTGACA

13. Promoters of Transcription
Eukaryotic promoters
May be a purine or pyrimidine
Usually purine in prokaryotes
Multiple promoter sequence
Order of boxes
TATA Box (25 BP from Initiator)
CCAAT (CAAT BOX)
GGGCG (GC BOX)
NOTE: RNA Polymerase III promoters occur downstream of the startpoint!
Between and

14. Initiation of Transcription
Prokaryotes
 (sigma) factor
Allows RNA polymerase holoenzyme to attach to promotor sequences
Process
 (sigma) factor facilitates opening of DNA
Enzyme (holoenzyme) forms phosphodiester bond between first two bases
Elongation begins and after 10 nucelotides have been added,  factor is released
Released  factor can bind with another holoenzyme and initiate transcription at some other location

15. Initiation of Transcription
Eukaryotic Initiation Factors
Need four (4) factors to initiate transcription from a TATA Box region
TFIID
Binds directly to the TATA Box promoter region
TFIIA
Binds with TFIID that is already bound to TATA Box
TFIIB
Facilitates RNA Polymerase II binding to aggregate
TFIIE
Binds to preinitiation complex and triggers beginning of transcription at initiation point (start point)

16. Termination of Transcription
Prokaryotes
Factor-independent termination
Inverted repeat causing a loop
Loop area rich in GC pairs
Stability for slow down or stop
Stretch of Uracils (poly U) after loop region
Factor-dependent termination
Certain sequences act as termination sequences in presence of Rho-factor ().
Eukaryotes
Not much known
Transcription does continue several BP beyond pseudo-termination with Poly A (Tail region)

17. Post-transcriptional Processing in Eukaryotes
Formation of 5’ Cap
Cap 0 – methyl group
Cap 1 – two BP
Cap 2 – three BP
Poly adenylation (Tail)
Sliceosome
SNURPS
Intronic excision
Contiguous exons

18. Protein Synthesis (Translation)
Sequence of triplet code form AA sequence on RER
Genetic Code
mRNA (CODON)
Redundancy
64 possible codes
20 Aas
Stop Codons (UAA, UAG, UGA)

19. Activation of tRNAs Coupling of AAs to tRNA Aminoacyl-tRNA synthetases
Activate Amino Acids and facilitate attachment to 3’ end of tRNA
Associated with anti-codon
Wobble-base
Repair of improper associated tRNA

20. Eukaryotic Ribosomes 80s from a 60s and 40s subunits 60s subunit
A-site (Aminoacyl Site)
P-site (peptidyl transfer site)
T-site (Termination site)