Prokaryotic Transcription And Control Of Gene Expression Flashcards
What is the size of E.coli genome and how many genes
~4300 genes, 5Mb genome
How many proteins are made at a time in E.coli and what type are made at high and low levels
~3000
High levels= ribosomal proteins (10,000/cell)
Low levels= regulators of transcription (<10/cell)
What are the two classes of genes
Housekeeping/ constitutive- always expressed, amount not highly regulated
Inducible- expression is induced or repressed, highly regulated
What are the reasons why gene expression is regulated
To conserve resources
Respond to changes in the internal/ external environments (nutrients, building blocks, toxins)
Ordered development
How are translation and transcription coupled. What speed do they occur at and why are they coupled (only occurs in prokaryotes as they have no nucleus)
mRNAs are rapidly degraded (half-life is 2-3min) so transcription and translation are coupled and occur quickly to prevent mRNA degradation
Transcription= 40N/s
Translation= 15 aa/s (similar to above)
Which DNA stand is the coding strand and which is the template strand
5-3’ is coding
3-5’ is template= transcribed into mRNA
What enzyme transcribes DNA into RNA in prokaryotes
RNA polymerase- DNA dependent RNA polymerase
What is an operon
Set of co-transcribed genes under the control of a single promoter
Why are genes in operons co-regulated and what are their transcripts
Genes usually have related functions so can be co-regulated. Can also have unknown functions. Transcripts are polycistronic meaning multiple genes/ cistrons encode multiple proteins
What are non-coding RNA (give examples) and how do they function
Dont code for proteins but may be processed
Eg tRNA, ribosomal RNA, CRISPR RNA
Can function alone or as RNA-protein complexes like ribosomes
Often bind specifically to other RNAs or DNA by base pairing eg mRNA codon- tRNA anticodon
What does CRISPR stand for
Clusters of regulatory interspaced short palindromic repeats
What are the two parts to CRISPR
Immunisation process (adaptation) and interference process
How does the immunisation process in CRISPR work
foreign DNA inserted
CRISPR Cas proteins expressed from cas operon which recognise foreign DNA and cleave/ cut out a piece
Curt out DNA is inserted into gene encoding for CRISPR ncRNA leading to tags of foreign DNA with common pieces/ repeats between them
How does the interference process of CRISPR work
CRISPR repeat spaced ncRNA transcribed into pre-crRNA which is processed into mature crRNA
crRNA combine with cas proteins and used as guide by cas complex to inactivate corresponding nucleic acid
What are the three steps of transcription
Initiation, elongation, termination
What is a promoter region and how are they recognised
Where RNA polymerase binds (alpha helicies of sigma factor bind to DNA major groove)
What is a sigma factor and how do they allow recognition of different promotor regions
Protein subunit of RNA polymerase. There are alternative types which under different conditions, recognise different promotor sequences. They coordinate distantly located operons
What are the different subunits of RNA polymerase (RNAP) holoenzyme
Beta subunits which are catalytic
Sigma which provide specific binding to DNA
Alpha subunits which are for enzyme assembly, promotor recognition and bind some activators
aaBB’ is the core enzyme, sigma provides specificity
Also w in some RNAP so can be aaBB’w
Steps in transcription initiation and elongation
Non-specific binding of RNAP, moves to promotor
Sigma binds when promotor is recognised and causes closed complex
Open-promotor complex formed where strands are separated
Initiation of mRNA synthesis- almost always with a purine
Initiation can be abortive- releasing 2-9 oligonucleotide chains)
Elongation of non-abortive mRNA
Release of sigma as polymerase proceeds down template away from promotor
What are the coding and non-coding strands in transcription of prokaryotes
Sense (coding) strand
Antisense (noncoding/ template) strand
What is Rifampicin (RIF) and how does it work
Transcription inhibitor of tuberculosis protein. Resistance to this can arise
Binds to catalytic beta subunit of RNAP and stalls transcription (used with two or three other drugs for 6 months)
If resistance occurs, other drugs must be used instead
What are the two methods of transcription termination and a brief overview
Rho independent- most common, specific hairpin structure in RNA formed from complementary base pairing in the RNA and the poly-A tail creating a poly-U tail in RNA at the end
Rho dependent- Rho protein (hexomer) binds to RNA and stops transcription, is ATP dependent also
Features of RNA that differ to DNA and make it able to form a hairpin for transcription termination
RNA helix has hole down the middle
RNA has narrower and deeper major groove
Extra OH group
Same number of bonds between bases when they pair (A=U and C=-G)
Bases at the top of the loop (actual round part) do not pair together
What things does the regulation of transcription depend of
Promoter strength- degree of match, weak or strong for sigma factor
Presence of alternative sigma factors active under different conditions
Presence of transcription factors via positive or negative regulation (binding affinity of these can be modified allosterically or covalently
