Transcription I Flashcards
regulatory component of gene
controls rate/efficiency of transcription
strucutral component of gene
contains necesary information to make product
operon
multiple genes and corresponding regulatory sequences that work toward a specific biological function under the control of a single promoter
viral transcription
both strands of dsDNA can serve as templates for transcription; top template strand has is initially transcribed as a single large transcript and then processed to give different products
polycistronic mRNA
one RNA transcript codes for multiple gene products, not in eurkaryotes
monocistronic mRNA
one promoter and one gene product, seen in eukaryotes
eukaryotic RNA polymerase II
makes mRNA, requires Mg2+
eukaryotic RNA polymerase I
makes precursor products for the 5.8S, 18S and 28S rRNAs, requires Mg2+
eukaryotic RNA polymerase III
makes tRNA and precursor products for the 5S rRNA, requires Mg2+
conventions for transcriptions
reads template/noncoding/negative strand in the 3’ to 5’ direction to create an RNA transcript in the 5’ to 3’ direction; RNA transcript will read the same as the non-template/coding/positive strand (except T’s replaced with U’s)
prokaryotic RNA polymerase (holoenzyme)
2 alpha, 1 beta and 1 beta prime subunits synthesize RNA, an omega subunit, and sigma subunit binds to DNA
beta sites are believed to be the catalytically active regions, the omega unit is not required in vitro
sigma RNA polymerase subunit
varible region that binds to DNA first, aligns polymerase into the correct orientation, sigma units dissociates once transcription starts, sigma 70 is the most common sigma unit
Step 1: prokaryotic transcription–binding
- ) sigma 70 subunit of RNA polymerase recognizes the -35 to -10 region and forms the closed complex
- ) unwinds DNA in this area forming open complex, DNA fed through RNA polymerase at a 90 degree angle, which forces dsDNA to open
- ) sigma subunit release once transcription proceeds beyond the promoter region
Step 2: prokaryotic transcription–initiation
- ) purine usually added first
- ) subsequent nucleotides added and PPi released
**PPi NOT cleaved from initial purine**
Step 3: prokaryotic transcription–elongation
- ) occurs in the transcription bubble, RNA polymerase covers 35bps and has melted 18bp, and there is an 8bp DNA/RNA hybrid region
- ) region ahead of polymerase has positive supercoils and the region behind has negative supercoils, which can be relieved by topoisomerases
rho independent trancript termination
string of adenines after hairpin structure, only two hydrogen bonds between A and U so mRNA weakly hybridized to DNA; polymerase falls off
rho dependent transcript termination
hairpin structure causes polymerase to pause, rho protein then causes dissociation
sigma 70 subunit
has a specific consensus sequence in the -35 to -10 region, changes in this promoter sequence decrease polymerase affinity and thus rate of transcription
using different sigma factors
example is under heat stress, polymerase dissociates from sigma 70 and instead binds to sigma 32–binds to a different -10 to -35 consensus sequence
operator site (repressor bidning site)
downstream of promoter, repressor proteins bind here and either block polymerase movement or block the promoter
activator site
just upstream of the promoter, enhancer proteins can recruit polymerases and increase transcription
negative transcription regulation
bound repressor inhibits transcription; 1.) small signal moleclar binds to the repressor, causes conformation change in affinity of repressor for DNA, falls off and transcription starts OR 2.) signal molecular always bound to repressor, when concentation of signal molecule drops, repressor falls off, transcription starts
postive transcriptional regulation
bound activator facilitates transcription, same two scenarios as negative transcriptional regulation
lactose metabolsim
- ) lactose enters cells through galactosidase permease
- ) lactose broken down by beta galactosidase into galactose and glucose
OR
3.) isomerized into allolactose
lac repressor gene
located just upstream of lac operon has own promoter (P1) and contains O3 coding sequence, when no repressor, it is producted and binds to O1 and sequently O2 or O3–causes homodimerization, DNA looping and cessation of transcription
allolactose
binds to lac repressor, causes conformation change, and repressor disscociates from operator, NOT sufficient enough to cause transcription of the lac genes
cyclic AMP receptor protein (CRP)
bacteria only use alternative fuel source if glucose not available, when glucose low, cAMP high, cAMP binds to CRP, CRP goes to CRP activator site–acts as enhancer, increases the binding affinity for sigma 70 and RNA polymerase to allow transcription to start
ara operon
encodes genes B, A and D necessary to metabolize arabinose and under the control of the BAD promoter
two operons (araO1 and araO2) and an inhibitor site (araI)
inhibitor that binds to araI is araC; encoded upstream of the operon and has its own promoter (promoter C)
how does the ara operon function?
no repressor–promoter C recruits RNA polymerase and makes araC—>araC binds to araI—>other molecules of araC bind to araO and the molecules dimerize, preventing transcription
when arabinose is present, it binds to araC and converts it into an ACTIVATOR, araC dimerizes with arabinose rather than ara I
requires CRP to initiate transcription
trp operon
contains all enzymes necessary to synthesize tryptophan, regulated by trp attenuation and trp repression
trp operon mechanism of action
SUFFICIENT TRYPTOPHAN: ribosome translate sequence one, which contains (two Trp codons) and then sits on sequence two and blocks it before sequence 3 is transcribed
sequences 3 and 4 transcribed and they form hairpin structure, terminating transcription
INSUFFICIENT TRYPTOPHAN: ribosome pauses at sequence 1, sequences 2 and 3 form hairpin structure but this does NOT cease transcription
-10 promoter region sequence
TATAAT
-35 promoter region sequence
TTGACA
