RNA Metabolism Flashcards
What does the unit symbol “S” represent?
Svedberg Unit (sedimentation coefficient) is a measure of how fast a particle of a certain mass, size, and density will sediment in a centrifuge.
Is there an absolute relationship between the mass of a particle and the value of “S” that is measured?
No, there are also factors such as shape and density of the particle that affect the sedimentation coefficient. For example, a large less dense particle may sediment slower than a small more dense particle.
Transcription
is the process where DNA is converted to RNA in order to be used for protein synthesis.
mRNA
carries the genetic information from DNA to the ribosomes for protein synthesis.
transfer RNA (tRNA)
delivers amino acid to the ribosome.
ribosomal RNA (rRNA)
forms the catalytic core of the ribosome.
small nuclear RNA (snRNAs)
RNA that are involved in splicing.
guide RNA
bind to specific ribonucleoprotein complexes and facilitate catalysis.
XIST
is a long non-coding RNA that resemble an mRNA but does not code for protein.
Barr Body
dense inactive X-chromosome found in nuclei of somatic cells in female mammals.
RNA vs. DNA sugar
DNA: deoxyribose
RNA: ribose
RNA vs. DNA sugar conformation
DNA: deoxyribose favors C2’ endo conformation.
RNA: ribose favors C3’ endo conformation due to extra hydroxyl group.
RNA vs. DNA preferred form in double stranded form
DNA: B-form
RNA: A-form
What are the major secondary structures of RNA?
stem, hairpin loop, pseudoknot, bulge, internal loop, and multiloop.
Housekeeping Genes
genes that are expressed at relatively constant levels throughout the cell cycle and play a critical role in cellular metabolism.
Constitutive Expression
genes that are transcribed at constant levels throughout the life of the cell.
Transcriptome
is the sum total of all RNA molecules made by an organism.
Promoter
is a DNA sequence located upstream of a gene and controls initiation of transcription.
Positive Control Elements
binding sites for activator proteins.
Transcription Ground State
is the activity of promoters in the absence of specific regulatory sequences.
Prokaryotic Transcription Ground State
promoters and associated proteins are generally active even in the absence of activator proteins.
Genomes are compact and lack the complex regulatory mechanisms.
Eukaryotic Transcription Ground State
promoters and associated proteins are generally inactive in the absence of regulatory proteins.
Access to promoters is restricted by chromatin.
Transcriptome vs. Genome
Transcriptome more selective.
Entire genome is copied exactly once, different variations in the composition of the transcriptome.
Genome long lived, transcriptome short life.
Chemical Mechanism of RNA Biosynthesis
IMAGE IN STUDY GUIDE LOOK
What is the nucleophile in the RNA biosynthesis?
3’-OH of the growing RNA chain
What is the electrophile in the RNA biosynthesis?
first phosphate of incoming nucleotide.
What is the leaving group in the RNA biosynthesis?
Inorganic pyrophosphate
What is the energy that drives the reaction forward in RNA biosynthesis?
the hydrolysis of the high energy phosphate bonds on the incoming nucleotide.
Initiation Step of Transcription
- RNA Polymerase binds to promoter, this leads to the closed complex.
- Partial unwinding of DNA leads to the open complex.
- Conformational change allows transcription to start.
TATA BOX
regulates transcription of a nearby gene, is a conserved DNA sequence located at the promoter region of eukaryotic genes.
What does the TATA box bind to?
The TATA box binds to the TATA box binding protein to form pre-initiation complex.
What happens to genes that do not have a TATA box?
Uses a protein TAF to bring TBP to the DNA.
What happens to the sigma subunit during initiation?
sigma dissociates from RNA polymerase before elongation happens.
NusA binds competitively to the polymerase and remains bound until transcription is complete.
Elongation Step of Transcription
RNA polymerase moves away from the promoter and forms the elongation complex.
Does RNA Polymerase have high processivity?
Yes, a single RNA polymerase will transcribe an entire gene without dissociating.
What are the two mechanisms that RNA Polymerases use for proofreading?
Kinetic Proofreading and Pyrophosphorolysis.
What is kinetic proofreading?
The RNA polymerase moves much slower if wrong nucleotide has been incorporated into a base pair.
Nuc: water, Product: NMP
What is pyrophosphorolysis?
cleavage of the nucleotide
Nuc: PPi, Product: NTP
What are the two different types of termination in transcription?
Protein Independent Mechanism and Protein Dependent Mechanism
What is the protein independent mechanism?
Termination sequence includes a segment of nucleotides that are self complementary and will fold into a hairpin structure.
HAIRPIN and Weak A/U base pairs in RNA cause the DNA and RNA to fully dissociate.
What is the protein dependent mechanism?
involves the use of a rho protein.
A stem-loop causes RNA polymerase to pause. Rho protein catches up to the open complex and separates the RNA-DNA hybrid.
Why do we need a rho protein in protein dependent mechanism?
Termination site lacks the AAA which causes the RNA-DNA hybrid to be more tightly associated which requires the rho protein.
RNA Polymerase Enzyme: what is it composed of?
complex of proteins made of core enzyme and sigma factor.
Alpha subunit of RNA Polymerase
promote assembly and binding to upstream elements.
Beta subunit of RNA Polymerase
carries the active site of the enzyme.
Beta Prime subunit of RNA Polymerase
binding to DNA.
Sigma subunit of RNA Polymerase
binds to the promoter sequences
What is the motif of the sigma subunit?
helix-turn-helix motif
What is the difference between prokaryotic and eukaryotic RNA Polymerase?
Prokaryotes has a single RNA polymerase that transcribes all types of genes.
Eukaryotes have multiple RNA polymerases that are specialized for different types of genes.
In the absence of proofreading, how will the correct nucleotide be incorporated?
Active site of the RNA polymerase plays a role in proofreading. The active site allows one nucleotide at a time.
Only the correct nucleotide fits well in the binding pocket.
Conserved Sequences
sequences that are shared between many different transcripts.
Inverted Repeat Sequences
when the nucleotide sequence on one strand is the reverse complement of the sequence on the other strand.
What are transcription factors?
proteins that help to express a specific set of genes by assisting in the process of RNA polymerase binding to the DNA.
How are transcription factors named?
named by the RNA polymerase they are associated with.
What are activator proteins?
allow transcription to happen.
What are some of the ways in which the activator protein allows transcription to happen?
- favor opening of DNA.
- Induce allosteric changes in the RNA polymerase that favor transition to the open state.
- Binds to the RNA polymerase and the DNA at a site next to the promoter, increases affinity of the polymerase for the promoter.
What are repressor proteins?
prevent transcription from happening
What are the mechanisms in which repressor protein prevents transcription from happening?
- Blocks promoter accessibility: bind to repressor sequence that overlap the promoter sequence. Create steric block that prevents RNA Polymerase from binding.
- Allosteric Mechanism: they bind to sequences that are close but not overlapping the promoter. Prevents opening of the DNA required for transcription.
What are architectural regulator proteins?
proteins that bind to DNA and facilitate the process of looping.
What are enhancers?
proteins that bind to distal regulatory sites.
Insulators
limit the looping
What is the CTC binding protein?
structurally bringing everything together for transcription to happen.
RNA Polymerase II CTD
plays a crucial role in transcription, initiation, elongation, and mRNA processing.
What are the post-translational modifications and structural alterations that are used to regulate RNA metabolism through the tail sequence of RNA Polymerase II?
- 2nd and 5th serine subject to phosphorylation
- phosphorylation of tyrosine
- Cis/Trans isomerization of two prolines.
What is the mediator complex?
essential for transcriptional regulation in eukaryotic cells.
