“central dogma”
Gene
• a DNA segment having the all information necessary for the synthesis of a functional RNA or protein
• it includes not only the coding sequences but also regulatory regions
Prokaryotic gene structure
• Prokaryotic genes are generally seen as structures called “operon”
• Basic elements in operons:
– Promoter – Operator
– Transcription start site – Coding sequence
– Transcription termination region (terminator sequence)
Promoter
A region of DNA, located on the 5’ end of the gene, where RNA polymerase binds
to initiate transcription
Promoter indicates the starting point of transcription
Promoter structure
• -10 region (Pribnow box): TATAAT
– unwinding domain
• -35 region: TTGACA
– recognition domain
seperated by 16-19 base pairs
• 5-9 base pairs downstream of the -10 region
• Generally (>%90) a purin nucleotide (CAT)
Transcription start site
Coding sequence: generally more than one protein is coded (polycistronic)
Termination region: a G-C rich region followed by 4-8 U nucleotides
Regulator sequences
• Operator
• Upstream elements (UP elements)
• Enhancers
Operator: located on -5/+20 and binds repressor protein
UP elements: located at -30/-65
Enhancer: located at -80/-160 and activates transcription
Eukaryotic gene structure
- bacteria have only one RNA polymerase
- eukaryotes have at least three different RNA polymerases, each
responsible for tanscription of
distinct class of genes
• RNA polymerase I Class I genes
genes coding 5.8S, 18S ve 28S’lik rRNAs
• RNA polymerase II Class II genes
genes coding mRNAs
• RNA polymerase III Class III genes
genes coding tRNAs and 5S rRNA
Class II genes
• Promoter structure:
– TATA box (TATAAA): -25 – CAAT box: -75
– GC box (GGGCGG): -90 – Octamer (ATTTGCAT)
– Initiator (PyPyAN(T/A)PyPy)
Coding sequence is interrupted by non-coding sequences
Transcription
termination region is determined by
polyadenylation (polyA) signal
Regulator sequences
• Promoter proximal element (PPE)
• Enhancer
• the first transcription product is called as primary transcript (pre-mRNA)
• further processing of pre-mRNA is required to become mature and
functional mRNA
Posttranscriptional modifications
• 5’ capping
• splicing
• polyadenylation
mRNA synthesis
Prokaryotic RNA Polymerase
subunit
– scaffold
– promoter binding
ve ’ subunits
– catalytic center
(sigma) factor
– promoter specific binding
Prokaryotic RNA Polymerase
Prokaryotic RNA Polymerase
Holoenzyme (2’)
• Core enzyme (2’)
– consist of the subuints needed for the catalytic activity
• Sigma factor ()
– promoter specific binding
loose binding
closed binary complex
open binary complex
ternary complex
abortive initiation
promoter clearance
Termination
• independent termination (terminator sequence)
• rho () dependent termination
independent termination
(terminator sequence)
Rho ()
dependent
termination
Class II Gene Structure
• Eukaryotic RNA polymerases can not bind to promoter itself and require several initiation
factors (general transcription factors) for efficient and promoter-specific initiaiton of transcription
• RNA polymerase and general transcription factors, bound together at the promoter and poised for initiation, is called the preinitiaition complex
TFIID
TBP (TATA Binding Protein) TAFs (TBP Associated Factors)
Posttranscriptional modifications
• 5’ capping
• splicing
• polyadenylation
Capping
• the addition of a modified guanine base to the 5’ end of the RNA
– a phosphate group is removed from the 5’
end of the transcript
– GMP is added (guanylyltransferase) and then modified by the addition of a methyl group (methyltransferase)
splicing
Polyadenylation (PolyA tail)
addition of polyA tail to the 3’ end of mRNA
• mRNA is cleaved at 11-30 nucleotides downstream of polyA signal by;
– CPSF (cleavage/poliadenilasyon specificity factor) – CstF (cleavage stimulation factor)
– CFI ve CFII (cleavage factors)
• multiple adenine nucleotides ( 200 copies) are added by PolyA polymerase
