Explore Essential Molecular Biology at UCL. Dive into the intricate world of RNA, exploring detection, manipulation technologies, RNA processing, and transcription. Tailored for Year 2 students, these notes provide a humanized academic guide, enriching your understanding of RNA dynamics. Please not...
RNA – Summary
RNA Transcription
RNA transcription – summary (DNA RNA protein)
o Transcription
Copying of coding strand of DNA (sense strand) into an RNA molecule (transcript)
RNA-transcript = identical to coding strand – but uses uracil instead of thymine
DNA is transcribed to RNA – by RNA polymerase
Link nucleotides to form an RNA strand – using DNA strand as a template
o Phosphodiester bond between 3’ OH of growing RNA strand + 5’ phosphate
of incoming rNTP
o RNA molecules must be processed after transcription
Spliced have a 5’ cap and poly-A-tail on their heads
o Translation
mRNA sequence is read by a ribosome – to produce a protein
RNA polymerase
o Main enzyme involved in transcription
o Uses a single-stranded DNA template – to synthesise a complementary
strand of RNA
Builds an RNA strand in the 5’ to 3’ direction
Adding each new ribonucleotide (rNTP) to the 3’ end of
the strand (rNTP = ribonucleotide tri-phosphate)
o Prokaryotic vs eukaryotic RNA polymerases
Differences
Prokaryotes have 1 RNA polymerase
Eukaryotes have 3 RNA polymerase
Made up of different subunits
Similarities
RNA polymerase is a large multi-subunit molecule
Transcribes DNA to RNA without needing a primer
Has a highly conserved structure = core enzyme – responsible for process of
synthesis + relies on extra proteins
o Structure
Prokaryotic RNA polymerase
Smallest
Complex of 400kD
Made of 5 subunits
o 2 copies of α subunit – scaffolding
rule
o 1 β subunit = active site where
rNTPs bind
o 1 β’ subunit = DNA binding
o 1 ω subunit = unknown role
Subunits assemble to form a large complex with duel-like lobes – separated by a
cleft
o Enzyme active site
At the base of the cleft + accessible through opening in β’ subunit =
2 points of access to active site
In some bacteria – β and β’ subunits are produced as a single polynucleotide
Eukaryotic RNA polymerases
3 eukaryotic polymerases
o R1, R2, R3
, RNA – Summary
Contain more subunits than RNA polymerases
o Ranging from 12-17 – depending on polymerases (Polymerase II = 12)
Stages of transcription
o Initiation
RNA polymerase binds to promotor separates the DNA strands providing single-
stranded template needed for transcription
o Elongation
Anti-sense strand = template for RNA polymerase
RNA polymerase reads the template (non-coding strand) one base at a time –
building an RNA molecule using complementary nucleotides resulting in a chain
growing from 5’ to 3’
RNA transcript carries the same information as the non-template (sense) strand of DNA
Contains uracil (U) instead of thymine (T)
o Termination
Terminators = sequence of DNA that signals the RNA transcript is complete
Once transcribed = cause the transcript to be released from the RNA polymerase
Eukaryotic RNA modifications
o In bacteria
RNA transcripts can act as mRNAs (messenger RNAs) straight away
o In eukaryotes
Pre-mRNA = transcript of a protein-coding gene needs extra processing before translation
Have their ends modified – addition of a 5’ cap at the beginning + 3’ poly-A-tail at
the end = increases the stability of mRNA
Undergo splicing – introns are removed + exons are joined back together
o Gives the mRNA its correct sequence
Promotor recognition – differs in prokaryotes and eukaryotes
o Locating promotor
RNA polymerase cannot locate the start of transcription (+1) on their own
Additional proteins guide core enzyme complex to promotor region
Promotor = sequence of DNA adjacent to transcription start site near the beginning
of the gene where RNA polymerase binds – specific to each gene
Prokaryotes = sigma factor
Eukaryotes = transcription factor
o In prokaryotes – sigma factor
RNA polymerase needs sigma factor (enzyme) = directly contacts promotor sequence –
ensuring correct start for transcription
Sigma factor subunit + RNA polymerase = holoenzyme
o Sigma factor binds to specific sequences upstream of transcription site –
guiding the RNA polymerase to the promotor
Bacteria have a variety of sigma factors = recognise different
promotor sequences – allowing cell to alter gene expression
Sigma 70 = common factor in E.coli – responsible for the
initiation of most genes
Bacterial promotors
Contain 2 key recognition elements
o Conserved sequence at -35bp
o Conserved sequence at -10bp
Spatiotemporal gene expression
o There are differences in spacing between conserved sequences – affecting
the binding affinity of different sigma factors
Variations contribute to the spatiotemporal gene expression
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