RNA and Protein Synthesis, Secretion and Degradation Flashcards
Some of the information in the slides is present in other lectures! Refer to DNA Replication Lecture for repeat items.
_________ is used in transcription. How does it work?
RNA polymerase.
Reads DNA and transcribes it to RNA
Prokaryotes have a _______ RNA polymerase while Eukaryotes have ____________.
single, three
Eukaryotic RNA polymerases recognize ______________.
Different types of promoters
What are the three eukaryotic RNA polymerases?
RNA I , II and III
RNA I transcribes_________
rRNA genes
RNA II transcribes _________
Protein-coding genes
RNA III transcribes ______.
tRNA genes, rRNA genes
Which eukaryotic RNA polymerase makes up the highest percent ?
RNA polymerase 1 (75%)
Transcription is the copying of a DNA template ______ direction to synthesizes a ______ RNA molecule.
Is a primer required for RNA transcription?
Transcription is the copying of a DNA template in the 3’-5’ direction to synthesizes a 5’-3’ RNA molecule.
Primer is NOT required.
What nucleotides are used in RNA transcription?
AUGC
What allows for the production of a phosphodiester bond between two nucleotides ?
The incoming nucleotide cleaves two phosphates to create pyrophosphate. This releasing energy drives the reaction to allow the remaining phosphate on the 5’ carbon of the incoming nucleotide to bond with the 3’ -OH of the existing nucleotide.
RNA Polymerase Features
Synthesizes RNA from ____ in _____ direction.
Uses ______ as substrate
Require ___ template
Does NOT require _____.
DNA in a 5’-3’
ribonucleoside triphosphate (rNTPs) (AUGC- TP)
DNA
primer
How does RNA transcription work?
Helicase will first break hydrogen bonds between the double-stranded DNA.
RNA polymerase, which does not require primer will go ahead and use one of the strands to create a new RNA strand in the 5 to 3 prime direction.
As the RNA displaces, the DNA helix reforms.
The RNA strand developed will be the same as the ______ strand of DNA since the RNA was made __________ to the _______.
The only exception is that all _____ will be replaced with _______.
Coding
complementary, template
Thymines with Uracils
How does RNA polymerase recognize specific genes?
What two things does it determine?
The promoter is an ASYMMETRICAL DNA sequence that determines where transcription of a gene my RNA polymerase that is upstream of the RNA-coding sequence.
- Which DNA strand will be the template ?
- Where on the DNA strand will the RNA polymerase start ?
What stops RNA transcription?
Terminator sequences on DNA
What are transcription factors?
Proteins that bind promoter regions and recruit RNA polymerase to specific genes.
Describe the role of transcription factors in relation to the insulin gene in B-Pancreatic cells and the Neuron
Both pancreatic cells and the neuron contain the insulin gene. However the insulin gene is turned off in the neuron and is transcribed in beta pancreatic cells. This is because beta pancreatic cells have cells type specific transcription factors bound to the promoter of the insulin gene that allow it to be turned on all the time.
Describe the steps of transcription
First a transcription factor recognizes a promoter and guides are in a polymerase to the site.
RNA polymerase reads the promoter to verify that it is a true region, but does NOT transcribe the promoter region at all.
After verifying that, it is a region that needs to be coded, RNA polymerase escapes the promoter. The transcription factors is released, and RNA polymerase proceeds to transcribe the necessary gene.
After creating the RNA strand RNA polymerase dissociate from DNA once encounters a stop sequence.
RNA polymerase II contains _____ and ______ which _____________ respectively.
exons and introns
Coding and non-transcribed sequences
Why is the readied RNA product not ready for translation?
What has to be done in order for it to be ready?
It contains introns which are unnecessary.
Protein modifications of a 5’ Guanosine cap and a 3’ poly-A tail as well as removal of introns is necessary.
What is the 5’ Guanosine Cap ?
The 5’ Guanosine Cap is an important post translational modification that protects transcript from degradation.
What is a poly-A tail?
It is a 3’ end post-translational modification that marks transcription termination, prevents RNA degradaion and allows transcript to localize to the cytoplasm.
Which post translational modification allows RNA transcript to localize to cytoplasm?
Poly-A tail
How are introns removed?
Introns are removed through splicing.
Why is splicing important for the creation of proteins?
Aside from removing the introns, splicing allows for multiple different types of proteins to be generated from one RNA transcript.
Different exons can also be removed by a splicing protein, allowing for a different protein, to be produced.
rRNA is synthesized by ________ in the _______.
RNA polymerase I in the nucleolus
What are the two things that make up a ribosome?
rRNA + protein
Segments of _______ are transported to the _______ to become _________ of the ribosome.
rRNA, cytoplasm
Small and large subunits
tRNA is synthesized by ______ in the _____ and then transported to the _______.
RNA pol III , nucleus, cytoplasm
tRNA brings _____ to the _______. It recognizes the sequence via ______ which are ________.
amino acids, mRNA
anticodons, three bases that are complementary to the mRNA being read.
The anticodon is a PART of tRNA
In prokaryotes rRNA is produced as a _________ that is cleaved to form ___________. Cleavage is catalyzed by ________.
_______ is also cleaved from a larger transcript.
a single long transcript , ribosomal RNA subunits, RNaseP
tRNA
Translation is _______ to _______. It requires a _________.
mRNA to AA
a LOT of energy
tRNA will have an amino acid attached to the ________.
3’ end
What is the initiation codon?
AUG (Methionine)
What are non-sense codons?
Stop signals
Describe the steps of reading the genetic code.
- mRNA is read 5’-3’
- Ribosome scans mRNA for AUG codon.
- ORF (Open reading frame) of RNA is translated until STOP codon is reached.
What is the ORF?
Start Codon , Sequence, Stop Codon.
Protein Synthesis requires 5 steps what are they?
- Attachment of AA to tRNA
- Initiation of protein synthesis
- Elongation of polypeptide
- Termination and release of protein
- Folding and Processing of Protein
How do AA’s attach to the _____ end of tRNA?
3’
There are 20 aminoacyl tRNA synthetases. Each synthetase recognizes
- The correct Amino Acid
- The correct set of tRNAs that can carry that amino acid based on a specific set of nucleotides on the tRNA (anticodon sequence)
An ester bond is formed .
What bond is formed between an AA and tRNA ?
Ester
How does mRNA get read by the _______ ?
The large subunit and small subunit of a ribosome creates 3 sites.
A - Aminoacyl
P- Peptidyl
E- Exit
The A site is most frequently occupied by aminoacyl-tRNA
The P site is occupied by peptidyl -tRNA which carries the growing peptide chain.
The E site hold tRNA on transit out the ribosome.
Describe initiation mRNA translation in the ribosome.
First, the small sub unit of the ribosome binds to the mRNA, the five prime cap on mRNA guides ribosomes to align with the mRNA in eukaryotes.
The code is identified, and the methionine tRNA is based paired to its codon.
The large subunit aligned itself to the tRNA molecule at the P site and forms a complex with the small sub unit. This completes initiation.
What is the first amino acid in prokaryotes?
Formyl-methionine
Where does the the first codon , ______, get added in the ribosome?
P-site, MET (AUG)
Describe elongation of the polypeptide once translation has started in the ribosome (after initiation)
After initiation, the methionine amino acid is in the P site of the ribosome
Amino acyl t-RNA will bind to the A-site if it brings the correct complementary tRNA to the mRNA being read. If not, it will leave and bring a new codon to try.
A peptide bond is formed between the AA in the P-site and the AA now in the A-site via peptidyl transferase.
The ribosome will translocate and move the growing chain to the P to allow for the next charged tRNA to align with mRNA in the A site of the ribosome.
The tRNA in the P-site will exit UNCHARGED.
What is peptidyl transferase?
It is a catalytic RNA component of the ribosome that carried out the formation of peptide bonds between amino acids in the P and A site of the ribosome.
Describe termination of mRNA translation
Elongation continues until the ribosome encounters one of the three stop codons in the mRNA, causing polypeptides synthesis to stop.
The stop codon in the mRNA is recognized by proteins, called release factors. Release factors buying to ribosomes and the bond between the peptide and the tRNA.
The protein is released and the ribosome dissociates from mRNA
Is there tRNA for stop codon?
NO
Many ribosomes can simultaneously translate a single mRNA to form a _________.
Polyribosome
In eukaryotes, protein synthesis occurs in the _________.
cytoplasm
In bacteria, translation and transcription are __________. Most protein synthesis occurs at ________.
linked, bacterial chromosome
Translation begins while mRNA is being transcribed and mRNA is short lived.
How do antibiotics work?
Antibiotics work by interfering with transcription and translation of bacterial proteins. They can bind to RNA polymerase
How does antibiotic resistance develop?
Bacteria can break down the drug via enzymes.
They can produce pumps to release the antibiotic out of the bacteria.
Tetracyclines bind to __________ to block __________.
Small ribosomal subunitl binding of AminoAcyl tRNA to the ribosome.
Describe folding and processing of proteins
AA will determine the way proteins fold into a 3D structure.
Proteins synthesizes on polysomes in cytoplasm remain ______.
Proteins synthesizes on RER can ________
there
be a secretory or membrane protein or be used in a lysosome
How do proteins know where to go?
They have signal sequences that are 15-20 aa long that dictate where they need to go. This sequence does get removed.
The hydrophobicity and helical conformation of the signal sequence helps traffic the protein.
How does the protein get transported?
The Golgi Complex modifies proteins via formation of disulfide bond, glycosylation or modification of carbohydrate chains.
They are then sorted into secretory or transport vesicle which bud from the trans face of Golgi
If the protein doesn’t fold correctly, what happens?
The protein can be destroyed.
Protein Synthesis can slow down
Attempt to refold protein
Cell Death
All occurs in ER
Examples of protein misfolding diseases
Alzheimers
How are proteins eliminated if they are done with use.
Most proteins are eliminated via Ubiquitin Proteasome System.
The rest are subject to the autophage/lysosome pathway.
Ubiquitin Proteasome System
76 aa size protein marks proteins to be degraded by proteases contained within structures called proteasomes.
3 proteins are required to tag the protein for destruction
- Ubiquitin Activating Enzyme (E1)
- Ubiquitin Conjugating Enzyme (E2)
- Ubiquitin Protein Ligase (E3)
A tail of ubiquitins is needed to activate protease activity on protein.
The ubiquitin system has been being considered to target ________ disease which can cause _______.
_________ is a proteasome inhibitor drug. How does it work?
Periodontal, inflammation
Proteasome
It inhibits the NFKB inflammatory pathway by preventing degradation of repressor protein.
