Exam 2 Flashcards
making a copy of the DNA
replication
synthesis of RNA from DNA
transcription
synthesis of proteins from mRNA
translation
How do bacteria make more cells? How do eukaryotes go through cells?
Bacteria - replicating and dividing
Eukaryotes - mitosis and meiosis
Where are linear chromosomes found?
Eukaryotic cells and some viruses
What is the shape of bacterial and archaeal chromosomes?
Circular
How many chromosomes are found in a bacterial cell? How many are found in eukarya?
Bacteria - one
Eukarya - 46
How many base pairs are in archaea, and what genes are included in the chromosomes?
4.6 million base pairs
Includes necessary genes for everyday circumstances
How many base pairs are in bacteria, and what genes are included in the chromosome?
94 thousand base pairs
There are no housekeeping genes, just genes for special circumstances
What are the main parts of the structure of DNA?
- Hydrogen bonds connecting nucleotides
- antiparallel strands
- sugar phosphate backbone/phosphodiester bonds
Why are hydrogen bonds good for connecting nucleotides?
They are strong but can come apart in heat which is useful to translation and transcription
What is the difference in structure between DNA and RNA
RNA: 2’ Carbon connects to an oxygen
What carbons are involved in connecting each deoxyribose to phosphate in DNA?
5’ and 3’
How is eukarya DNA stored?
DNA is coiled around histones because the cell only needs access to certain parts of the DNA at a time.
How is archaea DNA stored?
Some archaea use histone proteins, but most supercoil their DNA
How is bacterial DNA stored?
DNA is supercoiled which allows the cell to access only the needed DNA/loops
What do bacteria and archaea use to supercoil DNA?
DNA gyrase
What are the size dimensions of an E. coli chromosome?
- 6 million base pairs, 0.34 nm per base pair
1. 56 mm long chromosome packs into a 2 micrometer by 0.8 micrometer cell
Where are bacterial and archaea chromosomes located?
nucleoid
Where are the eukaryotic chromosomes located?
nucleus
Status of plasmids in bacteria and archaea
Extrachromosomal plasmids
Status of plasmids in eukarya
Plasmids are rare; mitochondrion and chloroplast have their own DNA
Which domain has chromosomes with transposable elements?
all
The largest plasmids (Archaea) are so large they might turn out to be small chromosomes
Halobacterium and Halococcus
RNA translated into protein
mRNA
RNA that carries amino acids to be put into proteins
tRNA
brings together mRNA and tRNAs for protein synthesis
rRNA
DNA sequence upstream of transcription start site
promoter
adds ribonucleotides complementary to the DNA molecule
RNA polymerase
sequence where transcription stops
terminator
How many genes are transcribed at a time in eukaryotes
one
takes the introns out of the strand so that the eons can exit the nucleus and have a cap/tail added in the cytoplasm
splisozome
line on RNA polymerase in eukaryotes to help bind to promoter
transcription factors
what binds transcription factors in eukaryotes
TATA box
in this, TFB lines RNA polymerase at the starts. It also has a TATA box, and TBP is the TATA binding protein
Archaea
In only bacteria, this binds to promoter then RNA polymerase starts transcription
Sigma factor
What does RNA polymerase read when transcribing DNA
Reads the 3 to 5 strand
Makes a 5 to 3 strand
How many RNA polymerases can read the DNA at once in bacterial transcription
there can be multiple RNA polymerases to amplify the signal
Describe bacterial transcription
Sigma recognizes promoter and initiation site. Transcription begins; sigma released. RNA chain grows. Termination site reached; chain growth stops. Polymerase and RNA released.
What is the difference in genome of a prokaryote compared to a eukaryote
Prokaryotes
- do not have intervening sequence
- no nucleus so no cap or tail
- the RNA is already mature
What makes prokaryotes able to cotranscribed multiple genes
polycistronic mRNA
Two ways to terminate transcription in bacteria
Rho-dependent & Intrinsic/Hairpin
How does Rho-dependent termination work in bacteria
Rho binds to RNA moving towards RNA polymerase - DNA complex. Rho removes RNA polymerase when RNA polymerase reaches the Rho-dependent termination site
How do intrinsic terminators work in bacteria
Inverted repeats in DNA sequence forming a stem-loop (hairpin) structure after transcription
How does termination work in eukarya and archaea
AAUAA signals endonuclease cleavage
RNA polymerase of bacteria
one with 4 subunits
RNA polymerase of archaea
one with 8 subunits similar to Eukarya RNA polymerase II
RNA polymerase of eukarya
3, each with over 12 subunits, RNA polymerase II transcribes mRNA
Promoter Recognition Sequences in Bacteria
- 35 sequence (TTGACA)
- 10 sequence (Pribnow box, TATAAT)
Only exist in prokaryotes. Several genes cotranscribed. Encode proteins or rRNA that are used together.
Operon
What types of operons are in E. coli?
amino acid and sugar operons to make amino acids and break down sugars, respectively
involves enzyme repression and induction
negative control of transcription
occurs when a sufficient product is present to stop synthesis of enzymes no longer needed
enzyme repression
Arg operon is an example of enzyme repression. How does it work?
In the absence of arginine, the repressor is not bound to the operator.
In the presence of arginine, arginine binds to the repressor allowing the repressor to bind to the operator to block transcription.
Arginine acts as a corepressor
occurs when the substrate is present to make enzymes needed to use substrate
enzyme induction
Lac operon is an example of enzyme induction. How does it work?
Int he absence of lactose, the lac repressor binds to the operator to block transcription. In the presence of lactose, the allolactose binds to the repressor to move the repressor off of the operator to allow transcription
activation of transcription controlled by binding of activator protein when inducer is present
positive control of transcription
Why is catabolite repression an example of positive control of transcription
It regulates if an activator protein can bind or not, which is why it’s under positive control. Glucose, lactose, maltose are all catabolites that break down molecules to make ATP.
Describe the “glucose effect” of catabolite repression
Glucose can repress another catabolite’s use. When the cells run out of glucose they switch, transcribe the lac operon, and start growing on the lactose. Always use glucose first because it is the simplest/easiest. Diauxic growth - when it finishes using glucose, the cell stops growing and looks for something available like lactose.
Overall regulation of the lac system
Activation: Binding of CRP to cAMP recruits RNA polymerase to start transcription.
Repression: The inducer (allolactose) separates from the repressor to activate it. The active repress binds to the operator and blocks transcription
Regulation of the lac system
POSITIVE CONTROL: In the absence of glucose, there is sufficient cAMP to bind to CRP. CRP is an activator protein and when bound by cAMP can bind to the activator binding site in the promoter of the lac operon.
NEGATIVE CONTROL: In the presence of lactose, lactose is taken into the cell and converted to allolactose. Allolactose binds to the lac repressor removing the lac repressor from the lac operator.
Regulation of the mal operon - positive control
In the absence of maltose, there is no activation of the mal operon.
Activation - in the presence of maltose (inducer), maltose binds to the maltose activator protein to bind to the activator binding site to activate transcription.
more than one operon under the control of a single regulatory protein (ex. Maltose and arginine in E. coli)
regulon
Control of transcription in Archaea by repressor proteins like NrpR
NrpR blocks TFB and TBP binding - no transcription.
NrpR binds alpha-ketoglutarate. This releases the NrpR from the DNA.
When NrpR is released TBP and TFP can bind - transcription proceeds
How does the 2 component regulatory system work?
The sensor kinase detects the environmental signal and autophosphorylates. The phosphoric group on the sensor kinase is then transferred to a response regulator that can bind to DNA and affect transcription
How does the multi component phosphorylation transfer system regulate sporulation
Sensor kinases recognize signals and become autophosphorylated. Changes phosphorylation of sporulation factors in the cell. Sigma factor is inactive when bound to a sporulation factor. Signal from endospore activates sigma factor, early endospore genes are transcribed. Signal from mother cell triggers synthesis of sigma factor in endospore and pro-sigmaK in the mother cell. Signal from endospore activates sigma factor.
mRNA sequence of three nucleotides
Codon
What does AUG code for in bacteria
formyl methionine
What does AUG code for in archaea and eukarya
methionine
Where is the open reading frame
from start codon to stop codon
What is the region before the start codon
5’ untranslated region
How many total codons exist?
64
What decides where a peptide can be in a cell?
Amino acids decide based on their characteristics (acid, basic, polar, non polar..)
What is the alternate amino acid that UGA codes for if there is a recognition sequence downstream of the mRNAt that forms a stem loop
selenocysteine
What is the alternate amino acid UAG can code for
pyrrolysine
Attached to the amino acid corresponding to the codon. Has anticodon sequence that temporarily base pairs with mRNA codon during translation
Aminoacyl Transfer RNA (tRNA)
Brings together mRNA and tRNA for protein synthesis
Ribosome
How does the ribosome move along the mRNA
one codon at a time
Overall size of the ribosome in bacteria and archaea
70S
Overall size of the ribosome in eukarya
80S
Ribosomal rRNA in bacteria and archaea
16S
23S and 5S
Ribosomal rRNA in eukarya
18S
28S, 5.8S, 5S
Total ribosomal proteins in bacteria? Archaea? Eukarya?
Bacteria: 56
Archaea: 67
Eukarya: 78
How many proteins are common to all domains
34
How many proteins are unique to bacteria
22
How many proteins are unique to archaea and eukarya
33 proteins are common too archaea and eukarya
11 proteins are unique to eukarya
Explain translation (APE)
Acceptance Site
P site: where the tRNA that was used will be so they can attach the first amino acid to the next one
Exit Site: empty tRNA leaves through here
Release factor comes in and dislodges the ribosome at a stop codon
Helps add the large subunit covalently to the small subunit
GTP
What is required to move each tRNA
i molecule of GTP
Translation in a prokaryotic cell
1 or more genes to 1 mRNA to one protein per gene in DNA. Each region is translated to create one polypeptide per coding region.
Translation in a eukaryotic cell
1 gene to 1 mRNA to 1 protein
several ribosomes can translate a single mRNA molecule simultaneously
polysome
Cells of this species can divide in six minutes because transcription and translation happen back to back
Vibrio
What is the main benefit of the quickness of division in vibrio cells
allows cell to respond to changes in the environment very fast
Induced mutations
Physical and Chemical
Spontaneous mutations
Errors by DNA polymerase in DNA replication. No control.
penetrates tissues, causes formation of ions that can break covalent bonds
ionizing radiation
low levels of ionizing radiation create
point mutations
high levels of ionizing radiation create
large chromosomal mutations
Difference between gamma and UV radiation
Gamma: will kill you
UV: will only affect your skin tissues
can survive exposure to gamma radiation that would kill humans
Deinococcus radiodurans
is found in soils and seeps up. If it is released into the air, it gets diluted. Responsible for 20,000 lung cancer deaths each year
Radon
Common intercalating agent used to stain DNA in gel electrophoresis. When bound to DNA, it fluoresces orange under UV light
Ethidium Bromide
Insert between bases in one or both strands that cause the helix to relax; inserts space in the strand and causes a frame shift mutation.
intercalating agent
Allows us to determine if a chemical is a mutagen, carcinogen, etc.
Ames test
What is the procedure for doing the Ames test?
Bacteria is plated on agar plate, and a disc is put in the middle of the plate. No histidine (essential amino acid) is on the plate. Control only has the water on the disk but the test disk has the chemical.If the chemical is a mutagen there will be more revertants around the disk indicating the increase mutation rate
Wild-type salmonella strain that had a mutation making it capable of making histidine
His-Plus
a change in one or a few base pairs that can occur anywhere in the genome and can affect gene expression
point mutations
New codon results in an amino acid that can result in a faulty protein. Most common when the first nucleotide is the one altered, but also common when the 2nd nucleotide is altered
Missense
New codon that is a stop codon. Results in a truncated protein (shorter).
Nonsense
New codon, but it codes for the same amino acid. Most common when 3rd nucleotide is the one altered
Silent
Three types of base-pair substitutions
Missense, Nonsense, Silent
A shift in the open reading frame caused by insertions or deletions. In most cases it will affect the regions around insertion or deletion location.
Frame shift mutation
Shift in the +1 direction
Insertion
Shift in the -1 direction
Deletion
2 classes based on how the mutation affects phenotype
Forward & Reverse
Functional to nonfunctional/or new function
Forward mutation
Nonfunctional/new reverts back to functional
Reverse mutation
Wild-type is chemotactic (run and change direction in response to chemical). Mutants don’t tend to change direction in response to chemical
Vibrio anguillarum
Wild-type have a smooth appearance. The mutants are rough.
Mycobacterium smegmatis
Mobile genetic element. DNA segment that can move from one position to another in the genome by non homologous recombination. Cause mutation by insertional mutagenesis.
Transposable elements
2 forms of transposable elements
autonomous and nonatonomous
Has transposase gene so they can transpose by themselves. Insertion results in mutable allele that is unstable
Autonomous transposable elements
Cannot transpose by themselves because they lack a transposase gene. Insertions are stable because they can’t move.
Nonatonomous transposable elements
Transposable element insertion into reading frame of gene results in loss of function mutation, typically complete loss of function
null mutation
How would a transposable element insertion result in an altered gene expression?
Transposable element promoter affect nearby genes
Classes of Transposable Elements
Those that move as DNA
Those that move as RNA and then convert to DNA for integration
What types of transposable elements move as DNA?
Bacterial insertion sequences (IS) elements and transposons (Tn)
What types of transposable elements move as RNA?
Yeast (Ty) transposons
Characteristics of Insertion Sequences
IS1, IS2, etc.
768bp - 5000+ bp in length
Have inverted repeats at the ends
Contains transposase gene that codes for the transposase enzyme that IS1 needs to move
2 types of transposons (Tn)
Composite (Tn10) and Non-Composite (Tn3)
Contains antibiotic resistance genes. Have IS element at each end that contain transposase genes needed for Tn to move. Conservative transposition. Transpositions result in target site duplications on either side of the transposons.
Composite (Tn10)
Contains antibiotic resistance genes. Contain transposase gene. No IS elements. Replication transpositions (makes copy of itself, keeps doubling). . Transposon results in target site duplication on either side of the transposon.
Non-Composite (Tn3)
Ty that codes for gag structural protein
TyA
Ty that codes for pol polyprotein
TyB
Progenitors of retroviruses like HIV
Transposons in yeast (Ty)
Replication error rates for humans
10^-5
Replication error rates for bacteria
10^-6
DNA virus replication error rates
10^-4
RNA virus replication error rates
10^-3
Mechanism of the SOS Response
DNA damage activates RecA which activates LexA protease activity. (The repressor LexA is self-cleaved, degrading it allows for transcription of many genes including several DNA polymerases needed to repair the damage.)
When the damage is repaired rica is inactivated and newly made LexA represses DNA repair genes
