Flashcards in Chapter 7: From DNA to Protein Deck (92):
What is the difference between DNA and RNA?
The 3' is the same. They both have OH, but at the 2' DNA is H only and RNA is OH
Also in RNA t becomes U
What can adopt a more complicated structure: DNA or RNA?
RNA. Because it is single stranded it can adopt more complicated shapes
What is the central dogma?
genetic information in the form of DNA is read through transcription and results in RNA and then by translation leads to protein.
What is more stable RNA or DNA more stable?
DNA is moe stable RNA is very sensitive to enzymatic cleavage. O on the 2' of DNA can react with many things
U makes it less stable than T
Is RNA single stranded or doubles stranded?
It is single stranded but often contains short stretches of nucleotides that can base-apri with complementary sequences found elsewhere on the same molecule.
This and non-conventional base paid interactions (A-G) allows RNA to fold into 3-dimensional structures determined by its sequences of nucleotides.
What is the non-template strand called?
The coding strand because it has the same sequence as the RNA
What is the end of the RNA molecule?
5' because it is the first to be synthesized. It is too the left. Moreover, a stand of RNA is transcribed 5' to 3'. The template strand will be 3' to 5'
How doe she cell know which section to synthesize?
gene regulatory elements within each gene determine that.
Error rate DNA vs RNA?
RNA is 10000x more error prone
What does RNA polymerase do?
Opens up the DNA for coding
How many RNA base are paired to DNA bases at any time in the RNA polymerase bubble?
Describe the structure of RNA polymerase?
It enzyme with an opening on one side where the nucleotides enter. 7 nucleotides are bound to DNA at any one time. The is a another opening where the RNA strand exits. Transcription begins in the 5' end.
Many RNA polymerases can work on a single strand of DNA
What is mRNA (messenger RNA)?
Provides code for proteins to be constructed outside of the nucleus
What is rRNA (ribosomal RNA)?
structural pieces of ribosomes
What is tRNA (transfer RNA)?
It transports amino acids to ribosomes during translation. Moreover, it helps mRNA in translation
What is snRNA (small nuclear RNA)?
works with proteins to process RNA. For example intron removal of exon splicing
Does gene expression always require translation?
No, is the process by which the information encoded in a DNA sequence is translated into a final product that has some impact on the cell. If the RNA molecule is the gene's final product than in this case gene expression does not require translation.
What is miRNA/siRNA (microRNA/small interfering RNA) ?
22 nucleotide sequences that regulate the translation of mRNA. Moreover, regulates gene expression/
How much of the DNA in our genomes encodes for protein?
5% This means that the bulk of RNA codes for some other what we traditionally think
What is a transcription factor?
Loose term for any protein required to initiate or regulate transcription.
What is the sigma factor?
It is a transcription factor in e coli. A subunit of RNA polymerase that recognizes the promoter of a gene for transcription to commence. When transcription begins sigma factor is released and the polymerase moves forward synthesizing RNA. This this connives until a terminator on the gene is reached. The polymerase and RNA strand are then released. The polymerase associates with another free sigma factor and the process begins again elsewhere.
In e coli there are roughly 7 different sigma factors and each one is responsible for recognizing the start of a particular group of affiliated processes. Eg, housekeeping, heat shock, nitrogen management.
Said another way related functions are all recognized by the same transcription factor.
What would happen without sigma factor?
RNA polymerase would transcribe at random
Where does transcription start?
At he +1 position. Before the that sigma factor of the polymerase recognize a sequence at -35 and then another sequences at -10. When it recognizes the sequences at -10 a bubble forms, the DNA strand are separated so transcription can begin once it falls off at +1.
When people refer to a gene sequence, what do they mean?
They mean the non-template strand because the non-template (sometimes called the coding strand) will have the same sequences as the RNA
Are promoter and terminator sequences included in the RNA?
The promoter is not, the terminator is.
What single termination of transcription?
First there is an inverted sequence with the ability to create a stem loop and then there is usually a sequence of TTTT. After the TTTT transcription stops
What is a stem loop?
It occurs when two regions of the same strand are complementary have nucleotide sequences when read in opposite directions. Once those base pairs bond and form a double helix, there are usually unpaired bases in the middle of the strand that create a loop at the end.
Sometimes called a hairpin loop
Can both side of DNA be transcribed?
Yes! The opposite strand can be transcribed since RNA polymerase always moves from the 3' to towards the 5' end, so the transcription will move towards the center of the gene.
What is the direction of movement of DNA strands of RNA polymerase?
Always form 3' to 5'. Which means it spit out the RNA strand 5' end first 3' end last.
What is the difference between eukaryotic RNA and prokaryotic RNA?
Prokaryotes have only one type of RNA polymerase and it transcribes mRNA, tRNA and rRNA.
Eukaryotes have three types of RNA polymerase I, II, III
Also, the only thing bacteria needs to initiate transcription is a sigma factor whereas eukaryotes require the assistance of many transcription factors which must assemble near the promoter cite before transcription can begin.
Finally, prokaryote genes are very close to each other, but eukaryotes can have up to 100,000 nucleotides pairs in between genes. This allows for long stretches of regulatory DNA sequences and can lead to very complex transactional regulation.
What does RNA polymerase I do?
transcribes 3 major rRNAs
What does RNA polymerase II do?
transcribes mRNA and some snRNA (most attention paid because it makes protein, making it the core to the central dogma)
What does RNA polymerase III do?
transcribes tRNA, one rRNA, and snRNA
What is TATA box?
A potential site where a transcription factor binds. There is a TATA binding protein (TDP) that binds here. It stradles the DNA and deforms the strand and indicates to the polymerase to bind there. Many other transcription factors needed to transcription bind here including protein that pull apart the DNA to create the transcription bubble.
Moreover, it provides a signal to the proteins to assemble at the transcription initiation site.
Where is the TATA box?
It is usually at -30 before +1 the transcription start site. Almost all mRNA genes have this sequence at they relative position.
How do the RNA nucleotides attach to the template strand?
What ends most mRNA in our body?
The poly A tail. Anywhere from 150-200 A's is added enzymatically after the RNA has been synthesized. These are on the 3' end. This tail is non-coding.
What added to the 5' front after transcription?
A methylated (CH3) guanine residue is added.
What does the poly-A tail and the guanine cap do for RNA?
Protein binding site for nuclear export (this is how it uses exporting machinery. Only protein with these structures can pas through the nuclear core. This prevents unprocessed RNA from being exported out of the nucleus. These structures stability, too.
What is the nucleolus?
A site inside the nucleus that is where ribosomal RNA are synthesized and combined with proteins to form ribosomes.
What are exons?
The coding regions of genes.
What are introns?
The non coding regions of genes
Do prokaryotes have introns?
No, their DNA is contiguous; it has no non-coding regions.
Besides adding the DNA cap and tail, what processing of RNA must happen in the nucleus?
The introns (non-coding DNA) must be removed.
Do proteins carry out splicing of RNA?
No, it is carried out by RNA molecules called snRNA (small nuclear RNA). These are packed with additional protein tot for small nuclear ribonucleo-proteins (snRNPs)
where are nucleotides added to RNA?
The 3' (OH) hydroxyl group.
What is NTP?
Nucleoside triphosphate. The generic name for ATP, GTP etc. Mentioned in the book at powering RNA polymerase
What is the difference at the 2' position on DNA and RNA?
That is where DNA has an H only and RNA has a hydroxyl group (OH). This difference makes a significant structural difference between DNA and RNA. It is why the RNA alpha helix is differently shaped.
In RNA as in DNA, which nucleotide pairs have three hydrogen bonds?
In RNA (somewhat similarly to DNA), which nucleotide paired have two hydrogen bonds?
What is the spliceosome?
Formed by snRNP's (small nuclear ribonucleic-proteins) they carry out the splicing out of introns. These snRNp's use complementary base pairing to attach to the intron A bunch of snRNPs converge around the intron turning it into a lariat that is then cut off at an adenosine. The tow strips are then rebounded by the splicesosome
How do splicesosomes know where to go to work?
There are specific sequences that indicate it.
What the benefits of the intron/exon arrangement of genes?
Important regulatory functionGenes can be spliced in different ways and will therefor build different proteins. It speeds up the emergence of new and useful proteins, which appear in early evolutionary history to have arisen by mixing and matching of different exons from preexisting genes.
What happens after each splice?
Exon junctions complexes are added at that site.
How did we crack the genetic code?
The same synthetic nucleotide, U for example, was fed into a cell free translation system that contained ribosomes tRNA, enzymes and other small molecules. The resulting polypeptide told researchers what was created with that amino acid.
Said more succinctly, a variety of polymers were made and then scientist saw what polypeptide they translated as.
Capture method: adding radiolabled amino acids to a soup of the cellular components necessary for synthesizing protein, then centrifuge to create a pellet. Finally, measure the level of radioactivity in the pellet. assuming you know the mRNA you stared with, the concentration of radioactivity in the pellet will tell you which amino acids they used. Moreoever, that radioactive amino acids were captured by the mRNA present
How is the genetic code written?
with the 5' primer terminal nucleotide to the left
How large is tRNA? What does it look like? How many tRNA genes do we have?
Its schematic shape is often referred to as a clover; its tertiary shape is more like a twisted L.
We have about 500 tRNA genes, which make 48 different tRNA.
What is a codon vs and anti-codon?
A codon is the three letter sequence of the RNA code that indicate which amino acid is to bind at given spot. The anticodon, which is a region of the tRNA binds complementarily to the codon.
Charts of nucleotide sequences and the amino acids they represent are codons.
What happens at the 3' end of the tRNA?
To the 3' OH group, the amino acid attaches. There is a highly conserved motif "CCA" to which the amino acid attaches. FYI, the 3' group is at he tip of the shorter arm
Where is the anticodon on the tRNA?
It is at the other end of the L from the 3' end that hold the amino acid.
What is aminoacyl-tRNA synthetases?
IT is the enzyme that chooses the correct amino acid to covalently attach/charge tRNA with the right amino acids. For most organisms there is a synthetases enzyme for each amino acid, so there are 20 synthetases in all. The enzyme is larger than the tRNA so it both reads the anticodon and attaches the amino acid.
Defects in this enzyme could lead to attaching the wrong enzyme to a tRNA. Many diseases are one wrong amino acid. So, attaching the correct amino to the correct tRNA is very important.
How are polypeptide chains built?
tRNA bind at the A site with the appropriate codon to the match the t-RNA's anticodon
What is the ribosome made of?
Two subunits. A large one with 49 proteins and 3 RNAs and a small one with 33 proteins and 1 RNA. Together is about 80 small proteins and 4 rRNAs.
The bulk of the ribosome is actually RNA. It was the largest structure every solved with x-ray crystallography.
Knowing the structure allows us to design antibiotics faster and better than people did in the past. The proteins are on the surface, their main role is to stabilize the rRNA core. It is one of the only really large cellular structures that is not protein.
The large subunit is on top and its movement and it movement across the small subunit move the tRNA along.
Describe the 3 binding sites in the ribosome?
A, P, E--all make primarily of rRNA
A: The tRNA enters here and attaches to the polypeptide chain
P: When a tRNA moves to a P site, the amino acid is attached, but released from the tRNA the moment the amino acid on the new tRNA at the A site attaches to the p-site amino acid.
E: After the amino acid has been released, the empty tRNA ejected at the E site
Are the different types of ribosomes?
No, the protein produced depends on the mRNA tape fed in to the the ribosome. Ribosomes can make any protein.
We know the polarity of the mRNA, it is constructed 5' to 3', but what is the polarity of the polypeptide chain?
The n-terminal of the protein is made first, with each cycle adding one amino acid to the c-terminus.
What are ribozymes?
Ribosomes are a type of ribozymes. RNA that is capable of catalytic activity (said another way, they are able to serve as enzymes). There is good reason to suspect that RNA not protein were the first catalysts for living cells.
Where is the catalytic site of the ribosome?
The catalytic site is the site that brings the chemical partners together in just the right space to facilitate a chemical reaction that would normally not happen. The catalyst on ribosomes is the large rRNA subunit, specifically the peptidyl transferase site of the 23s rRNA . The residues on in the rRNA bring everything together in just the right way.
Specifically, the rRNA strand that makes up the active site is 23s rRNA of the large subunit. The catalytic site of this rRNA is called peptifyl transferase. It is is a highly structured pocket at orients the two reactant, the elongating peptide and the aatRNA
Why is the genetic code redundant?
More than one codon codes for a given amino acid
Are introns and old feature of genes?
How does mRNA exit the nucleus?
The NES tag attaches to the poly-A tail and binds to exportin and then taken out of the cell
What is a Aminoacyl-tRNA?
It is a tRNA that is charges with an amino acid. Sometimes written as aatRNA
How do aatRNA get to the catalytic site of the ribosome?
An aatRNA is bound to elongation factor TU. It enter the a-site of the ribosome, if the codon and anticodon are complementary (if not it is rejected and trial and error continues). It the aatRNA is a fit, the elongation factor hydrolyzes it GTP and dissociates from the tRNA. The ribosome catalyzes the forming of a new peptide bond and a conformation change occurs. It lurches forward closing its active site and moving the tRNA to the P and E site leaving the A site empty but unable to take new tRNAs. Then elongation factor F binds to the ribosome, hydrolyses its GTP, which causes the ribosome to return to its original conformation.
What are the two ingredients for polypeptide synthesis?
elongating polypeptide and charged tRNA
How does translocation of the tRNA happen and why is it necessary?
The tRNA must move along to make room for new aatRNA. It moves by conformational changes in the ribosome cause and
How fast are amino acids synthesized?
How many ribosomes in a eukaryotic cell?
2-20 amino acid synthesized per seconds.
several million in a eukaryotic cell
What does the small ribosomal subunit do?
It is on the bottom (according to the book's schematics) and it positions the mRNA so it can be read
How do cells make sure that synthesis begins with the right reading frame?
The cap helps the translation initiators bind to the small ribosomal subunit, methionine (Met) takes up the P-site. This translation apparatus binds to the mRNA, the RNA cap signals the start of the mRNA. The methionine charged t-RNA moves along the mRNA strand until it hits an AUG sequence. The translation initiation factors then leave and the large ribosomal subunit binds. A charged tRNA binds to the A site and translation is off an running. Met is always the first amino acids on proteins, if it doesn't have a role in the mature protein, there are proteases that will remove the methionine.
FYI, met internal to the protein use a different tRNA
What is the difference between prokaryotic and eukaryotic mRNA?
multiple gene can exists on a single strand of prokaryotic mRNA. Different ribosomes transcribe different sections. Said another way, a single strand can have multiple RNA binding sites of prokayotic mRNA.
Is there tRNA for the stop codons?
No, but there is a protein with a similar shape called a release factor. It fits onto the empty A-site when a stop codon is hit. The causes hydrolysis of water and facilitates the release of the new protein from the tRNA. The ribosomes machinery comes apart and will be used again elsewhere.
How is a cell able to make protein so fast?
Many ribosomes work on a single strand of mRNA. The mRNA curls up. That it is curled keeps the poly-a tail and the 5' cap sort of close to each other. Perhaps there is some communication between the two making sure the mRNA is intact so that effort isn't wasted transcribing a broken mRNA
How do some antibiotics work?
They interfere with ribosomes by interfering with the APE site, the confirmation of the peptidyl transferase so that tRNA cannot be added, block initiation, block transaction, block RNA polyerase.
How does tetracycline work?
it binds to the A-site of bacterial ribosomes
How does streptomycin work?
Prevents transition from initiation complex to chain elongation
How does chloramphenicol work?
blocks the peptidyl transferase reaction on ribosomes
How does cyclohgeximide work?
It blocks the translocation of tRNA in the ribosome
How does rifamycin work?
Binds to RNA polymerase and prevents it from working
What is the proteasome?
It is a large protein machine that breaks down proteins
What are proteases?
The degrade proteins to short peptides and then to amino acids. It does this proteins that are no longer woking or degraded, proteins whose lives must be kept short, misfolded proteins
How are proteins broken down?
By large protein machines called proteasomes, a central cylinder with an active site with proteases in the inner chamber. It is powered by ATP. There are stoppers on both ends the bind the protein destined for degradation. Once inside stuff is chopped into small pieces and the expelled out the top and the bottom.
How does a cell decide with protein to degrade?
Proteins marked for degradation have ubiquitin attached to them