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144 Cards in this Set

  • Front
  • Back
Differences between DNA and RNA?
DNA = thymine, deoxyribose; wider/shallower major groove, less rigid, easier to store/condense
RNA = uracil, ribose
What is a base + sugar called?
Base + sugar + phosphate?
Nucleoside
Nucleotide
Difference between ribose and deoxyribose?
OH at 2' C (usually another H)
Which of the nucleic acids is monocyclic? Bicyclic?
Monocyclic = Pyrimidines
Bicyclic = Purine
What is the significance of the 2'-OH group of RNA?
Facilitates auto-catalytic cleavage of (only) RNA; the RNA enzymes depend on this 2'-OH
Why does DNA have thymine instead of uracil?
Cytosine can easily degrade to uracil spontaneously so wouldn't be able to know if the uracil was supposed to be there or was there due to degradation (when repairing).
Why is DNA usually double-stranded?
- Stability
- Bases on inside of sugar-phosphate backbone are sequestered/protected
- Redundancy of information (2 copies)
Who utilizes the major and minor grooves of DNA?
Utilized by proteins
Why can RNA be single-stranded?
It is not responsible for carrying on genetic information so it can afford to be more transient.
Why is the outside of DNA negatively charged?
Phosphates
What are the different kinds of RNA? (Unstable, Stable, Other)
Unstable = mRNA (temporary messages)
Stable = rRNA (enzyme components) and tRNA (translator)
Other = ribozymes (have catalytic activity) and riboswitches (regulatory mechanisms)
Why does RNA fold to form secondary structures that partially line up?
Thermodynamically favorable to bind bases; not all line up because RNA is not totally redundant (only in certain portions)
What is a hairpin?
A type of RNA stem-loop
What is the origin of replication?
oriC - sequence at which relevant proteins bind to start replication
What recognizes oriC during replication?
DnaA:ATP complex which forms an open complex
What unwinds the DNA during replication?
DnaB aka Helicase
What adds RNA bases to prime the DNA for replication?
DnaG aka Primase
What are the four steps of DNA replication?
1) DnaA binds at oriC
2) DnaB / helicase runs along and opens up
3) DnaG / primase lays down short piece of DNA
4) DNA polymerase III copies DNA
In what direction does polymerase synthesize DNA in? What direction on the template strand is this movement?
5'-->3' (synthesis)
3'-->5' (template strand)
What removes the unwanted RNA from Primase and replaces with DNA bases?
DNA polymerase I
Which enzyme fills the breaks in the DNA (from the lagging strand replication)?
DNA Ligase
When does replication occur in prokaryotes?
It is always occurring in the midst of other cellular processes.
How long is the average amount of DNA? How long is a cell? How does this fit?
6 mm of DNA in 0.01 mm cell
Tightly packed genome, supercoiling, histone-like proteins
What is RNA polymerase made up of? What is it used for?
Subunits, key one is the sigma subunit.
It is used in transcription.
What are the four steps of transcription?
1) Recognition - binding to promoter by sigma subunit
2) Formation of open complex
3) Elongation
4) Termination - release at terminator
What recognizes the special promoter sequence during transcription?
Sigma subunit of the RNA Polymerase
Which is found in the transcript? The terminator, promoter or both?
Just the terminator.
What does simple termination of transcription involve?
G-C rich stem loop and series of A's which causes polymerase to fall off (independent of Rho protein)
What are the two kinds of termination of transcription?
Simple termination (Rho-independent)
Rho-dependent termination
What is an operon?
Multiple genes encoded by one promoter.
What are the similarities of eukaryotic and prokaryotic DNA replication?
- Protein complex opens the DNA (helicase)
- Bidirectional
- RNA primase involved
- Similar base insertion (leading strands/lagging strands)
What are the differences of eukaryotic and prokaryotic DNA replication?
- 10-100x slower in eukaryotes
- Numerous origins of replication (oriC) compared to only one
- Cell cycle dependent (no competition w/ transcription)
- One replication event at each origin (replication is not continuous in eukaryotes)
What are the differences in promoters between bacteria and eukarya/archaea?
Bacteria: -35 and -10 regions act as promoters; regulatory sequences are peripheral
Euk/Archaea: regulatory sequences come before TATA box promoter and help to identify TATA

"Bacterial promoters are recognized by polymerase primarily. Euks and Arch in general need the TATA binding promoters to bind and recruit polymerase."
What connects mRNA and polypeptides?
tRNA
What is the codon for starting? Stopping?
Start = AUG = met
Stop = UAA, UAG, UGA
If you change any single nucleotide in the DNA coding sequence of a genome it will not always lead to an amino acid change in a protein. Why?
- Due to Wobble Hypothesis, you can change in the 3rd position and still have the same A.A.
- Not all sequence is translated; therefore, mutation would be insignificant to protein product
What are ribosomes? What makes up the ribosome?
Machine where translation occurs
Made up of small (30S) and large (50S) subunits to make the ribosome (70S)
What enzyme is critical for putting the proper amino acid on the acceptor end of the correct tRNA?
Aminoacyl-tRNA synthease
What are the three steps of translation?
1) Initiation - brings two ribosomal subunits together placing the first a.a. in position
2) Elongation - adds a.a. as directed by mRNA
3) Termination
What are the steps of initiation of translation?
1) Initiation Factor 3 helps bind the 30S subunit to mRNA
2) IF1 binds to block the A site (E-P-A)
3) IF2 brings the tRNA to bind to the start codon
4) IF3 releases and the 50S subunit binds by the fmet-tRNA
5) GTP hydrolysis releases IF1 and IF2
What does GTP Hydrolysis do during translation? How much of the cell's energy is required for this?
GTP is hydrolyzed for each amino acid added
Uses 40% of the cells energy
Elongation occurs by random insertion, how can it be random? Why is this important?
If it doesn't fit, the tRNA is rejected. Concentration of different anticodons is important, may take a while if low conc.
When does termination of translation occur?
When a stop codon is in position A of the ribosome.
What does it mean that "transcription and translation are coupled"?
Ribosomes bind to the mRNA and begin translation protein before RNA polymerase has finished transcribing the mRNA.
What would happen if you deleted the terminator of a gene?
Its protein product will function almost normally.
What is a heritable change in the DNA sequence compared to a defined control?
Mutation
What is the strain to which all are compared (arbitrary, but critical)?
What is the organism (or strain) with a mutation?
Wild-type
Mutant
What is the DNA sequence of an organism?
Genotype
What is the behavior or appearance of an organism?
Phenotype
What are the three type of base substitutions? What do they imply?
Silent - doesn't change a.a. sequence
Missense - chage single a.a.
Nonsense - stop codon early
What are these examples of mutations called:
abcd klmn
a bcde bcde fghi
abcde xyz fghi
abcde ihfg jkl
Deletion
Duplication
Insertion
Inversion
In base substitutions, there are two kinds, what are they?
Transition: purine --> purine; pyrimidine --> pyrimidine
Transversion: purine --> pyrimidine; pyrimidine --> purine
What are three causes of mutations?
1) Errors in process (replication / recombination)
2) Environmental inputs (mutagens / carcinogens)
3) Human manipulation (molecular biology)
Where are frameshift mutations likely to occur?
In redundant runs of bases an extra base is inserted due to strand slippage in replication
What are three specific mechanisms that cause mutations?
- Electromagnetic radiation (UV light, X-rays, gamma rays)
- Transposons
- Oxidative damage
What are mobile, genetic elements which insert into DNA and disrupt genes which stops transcription and translation prematurely?
Transposons
What is the purpose of the Ames test? What is the basis of it?
To screen compounds for mutagenic properties; when not many cells are able to grow, add the mutagen and cells develop a mutation which helps them to grow.
What is mutation fixation a race between?
Repair mechanisms and DNA polymerase which is trying to replicate the DNA to make the mutation permanent; usually the repair mechanism wins.
What are five different mutation repair mechanisms?
- Mismatch repair
- Photo repair
- Methyl transferase
- Base excision repair, nucleotide excision repair
- Recombination repair (SOS repair)
How does 'mismatch repair' fix mutations?
- Recognition by MutS
- Excision between methylated bases (which marks old DNA) by MutL, MutH
- Repair by Pol I
How does photo repair fix mutations?
- Photolyase enzyme binds dimer
- Light photon activates reaction
- Base repaired
How does 'base excision repair' remove damaged bases?
- Multi-enzyme process which is similar to mismatch repair but differs in recognition.
- Sugar-Phosphate backbone is removed and gap repaired
What is the repair mechanism for serious DNA damage?
Recombination repair: at mutation, DNA polymerase skips over damaged region, gap is replaced by section from good chromosome; gap on good chromosome is replaced via DNA polymerase; original mutation fixed by other repair pathways.
A mutant cell will always have _____; does not necessarily have to have _____ or ______?
- a different DNA sequence than W-T
- different behavior than W-T
- different proteins than W-T
Why would it be almost impossible to discover an existing mutation in RNA polymerase?
Because RNA polymerase is an essential enzyme and it can't be deleted/altered.
A frameshift mutation in a coding sequence of a gene is most likely to do what?
Eliminate the function of a gene product.
What is the complete genetic content of a cell or organism including chromosomes, plasmids and prophages?
Genome
What are the mechanisms of vertical transfer?
By replication and cell division (parent to offspring)
What are the mechanisms of horizontal transfer?
Any route other than by reproduction; acquisition of new genes from another organism.
Is there a correlation between genome size and organismal complexity?
No - Humans have significantly smaller genomes than newts and lilies.
What do chromosomes code for?
Essential genes; can have more than one.
Plasmids can code for what?
Useful functions although others are not useful (may be important to live in certain environments, but not necessary for general survival or it'd be called a chromosome).
What is an integrated plasmid?
A plasmid that becomes integrated / a part of the chromosome but can still be removed and passed on.
How do plasmids replicate?
Use host replication machinery but must provide enough information in order to be sure the host can replicate.
What does the "addiction system" protect against?
Cells that don't receive the plasmid of interest are killed by Doc toxin which is neutral when paired with the plasmid.
What are the vehicles of horizontal gene transfer?
Plasmids which are acquired and/or lost by the cell; kept around by selective pressure
Where is most of the genomic information kept?
One circular chromosome (some have more than one, some have linear)
Many have plasmids
In the ampicillin and tetracycline resistant picture, how do ampicillin-only-resistant and tetracycline-only-resistant bacteria survive in the culture with both tet and amp?
Horizontal gene transfer leads to single colonies that are both "amp and tet resistant"
In order for DNA to be relevant it must be capable of what two things?
Transfer and Capture
In what ways can DNA be transferred?
- Transformation (uptake of naked DNA)
- Conjugation (movement of DNA involving cell to cell contact)
- Transduction (movement of DNA by virus)
What is homologous recombination used to repair?
Severely damaged DNA; also helps incorporate foreign DNA
What is the transfer of genetic material as free DNA from a donor cell to a recipient?
Transformation
What are two mechanisms of artificial transformation for species who can't perform natural transformation?
- CaCl2 treatment - Ca allows DNA to stick to cell; heat treatment permeabilizes cell
- Electroporation - electric current makes cell momentarily permeable
What is necessary for natural transformation of DNA into a new cell?
Translocasome which helps bind DNA and incorporate a strand; phase-dependent.
Which method of DNA transfer requires cell-to-cell contact?
Conjugation
What is a cell designated as when the chromosome has integrated the "F factor" plasmid (from conjugation with a F+ cell)?
Hfr, or high-frequency recombination, strain
What is the transfer of genetic information between cells through the mediation of a virus (phage) particle?
Transduction
What are the two main steps for transduction? What two kinds are there?
1) DNA packaged into protein coat
2) Injected into host cell with or without accompanying viral (phage) genes
- Generalized of Specialized
What DNA can be introduced via 'generalized transduction'? 'Specialized transduction'?
Generalized - any DNA can be introduced
Specialized - aka restricted - only closely linked genes can be transferred
What is the process of moving a short "----" sequence of DNA within or between DNA molecules? What are the two sub-types?
Transposition ("transposable")
- Composite
- Complex
What is the predominant method of genetic exchange?
Bacteriophage and viruses
What do viruses package their genomes inside of?
Capsids - protein coats/shells
Are virus genomes DNA or RNA? What information does it contain?
Can be DNA or RNA;
Genomes contain information for initiating and completing an infectious cycle
What does a virus do when it infects a host?
- Virus replicates inside host
- Parasitizes necessary host functions
- Directs synthesis of new virion components
What is the function of the capsid?
Protects genome from temperature, chemical, and physical damage.
What are the three capsid structures we addressed? Describe.
- Icosohedron - made of 20 identical triangular faces (capsomer)
- Filamentous - capsomers arranged as a hollow spiral
- Asymmetrical Virus Particles - for bigger viruses
What forms the envelope that sometimes forms to enclose the capsid of a virus? What is the space between the envelope and the capsid called? Function?
The cell membrane of the host.
Tegument (contains accessory proteins from virus that are necessary for synthesis)
A virus is metabolically _____, but not biochemically _____.
"inert"
"helpless"
What are the steps of the Lytic Cycle?
- Attachment
- Entry
- Replication
- Translation
- Assembly
- Release
How do viruses attach to hosts?
Randomly; viruses are non-motile and depend upon random collisions with host receptors, therefore, as conc. of viruses increases so does chance of infection
How do viruses recognize the host receptors? What are these receptors usually used for?
They encode surface proteins that recognize receptors; these receptors have other functions for the host (metabolite/ion transport, motility, environmental sensing, etc.)
How do viruses enter host bacterial cells?
When virus recognizes host's receptors it changes the viral capsid which causes the nucleic acids to be released into the cell for further development.
What can be translated to a protein? What can this be made from?
+RNA (+mRNA)
From: ±DNA, ±RNA, -RNA
How can RNA viruses produce the +mRNA they need for translation? What is the concern with this?
RNA-RNA polymerase that the virus must provide (hosts do not have this); RNA polymerases do not proof read, so there is a much higher error rate
When does the envelop get put on the virus?
During release from the host the membrane envelops the virus capsid.
What is a viroid?
Small, infectious, naked nucleic acid molecules that form plant pathogens.
What are prions?
Single small, infectious protein molecules that cause misfolded host proteins.
What is the process of the lysogenic cycle?
- Attachment
- Insertion of DNA
- Viral DNA integrated into host genome
- Host cell replicates to form more cells with viral DNA
- Can re-enter Lytic Cycle by excision of phage DNA
How does regulation occur in microbes?
In response to stimuli
Where does regulation occur?
- During transcription
- Post-transcription
- Post-translational
When does regulation during transcription usually occur? What two kinds of proteins can be involved?
During initiation:
- Repressor - which prevents RNA polymerase action OR
- Activator - which facilitates RNA polymerase action
Describe what happens during negative regulation during transcription.
- Repressor binds near promoter
- Binding of repressor blocks promoter
- RNA polymerase cannot transcribe operon
Describe what happens during positive regulation during transcription.
- Activator binds near promoter
- RNA polymerase binds at promoter
- Transcription proceeds
A part of the mRNA that binds metabolites to hide the ribosome binding site, making it inaccessible, are called?
Riboswitches
How do small RNAs regulate transcription?
Binds to sigma subunit and reduces transcription at some promoter.
If regulation occurs near the beginning it is what two things compared to if it were later in the process?
Slower but more efficient.
Since regulators occur in a population, are cells ever 100% on or off?
No, forms an equilibria where the majority of activators are either bound or unbound depending on the concentration of the effector.
What is being monitored in attenuation?
Status of translation which regulates transcription.
Due to attenuation, when there is a high amount of trp, there is ____ expression because ______.
When there is a low amount of trp, there is ____ expression because _____.
high trp = low expression because ribosome reads through regions 1&2.

low trp = high expression because ribosome pauses and only covers region 1
What are the three methods of post-translational regulation that we learned about?
- feedback inhibition - regulates biosynthetic pathways
- enzyme modification - add small molecule which changes protein's activity
- enzyme degradation - remove enzyme via proteases
In a catabolic pathway (breakdown), it is ___ when substrate is present and ___ when substrate is absent.
On
Off
In an anabolic pathway (synthesis), it is ___ when substrate is present and ___ when substrate is absent.
Off
On
Where are sensor kinases found? What do sensor kinases do?
- In cell membrane
- Sense environmental stimulus
- Phosphorylates itself
- Transfers phosphate to response regulator which either inhibits or stimulates gene expression at operator
In the Sensor-Kinase and Response-Regulator system how is the phosphate removed from the response regulator?
Phosphatase removes it.
What do phosphorelay systems, or regulatory cascades, allow for?
Allows sequential expression of genes (useful in development).
Sporulation involves complex control of over 200 genes, what mechanism regulates this expression?
Phosphorelay system, or regulatory cascade; requires sequential expression
What would be the advantage to having three different sensor kinases?
Each detects environmental signals and therefore can distinguish between actual stimulation and a fake one; wouldn't want to perform process inappropriately.
What do global transcription regulators affect?
Multiple genes in response to a given condition.
What is adaptation?
Ability to change behavior in response to demands placed on the individual by the environment.
What action can allow survival of microbes when they have been exposed to lethal doses?
Pre-exposure to stress at a sub-lethal dose allows the microbe to adapt to better defend itself in the future.
What kind of mechanisms work globally for regulation?
- Regulatory proteins
- Alternative sigma factors
- Small RNAs
What rescue repair system is induced when lots of DNA damage has occurred?
SOS: >30 genes involved in DNA repair
Where are small RNAs found?
The intergenic regions that encode small untranslated RNA that have various functions.
How can sigma subunits be used to regulate transcription?
There are various, alternative sigma factors that can be utilized to direct transcription of distinct sets of genes.
In the E. coli example of alternative sigma factors, how was the heat shock treated?
- Normal temps: secondary structure hides ribosome binding site
- Higher temps, 2' structure melts; sigma-H protein translated
- Along with RNA polymerase, the sigma-H factor binds to the "heat-shock genes" for expression
Genes of a global regulon involving a DNA binding protein are all likely to have what two things?
- Same DNA sequence around promoters
- Not same promoter sequence or same protein product
How do you determine the genes regulated in response to any shock?
Monitor the mRNA synthesis
How do scientists deduce the roles of certain genes?
- Start with a system (cell)
- Remove a component of the system (mutation)
- Observe the resulting perturbation of the system (growth phenotype)
What is the difference between a "screen" and a "selection"?
Screen - look at all cells and find the ones with the phenotype you are interested in
Selection - define a condition where only the cells with the phenotype of interest grow
Out of "screening" and "selecting", which is more powerful? Easier?
Selection is easier and more powerful.
Screening is harder and less powerful, but more applicable to more phenotypes.
What five things are needed, in general, for the basics of molecular biology?
- DNA
- restriction enzymes
- replicon
- polymerase
- ligase
Why is it okay to occasionally have mutations?
So much product, that most will be original.
What does Polymerase Chain Reaction do?
- Mimics replication with a few differences
- Makes more DNA
What two enzymes are critical for "cutting and pasting" DNA?
- Restriction enzymes for cutting at sticky or blunt ends
- Ligase for pasting