Glossary of Microbiology: Chapter 11

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Define Genome
Genome is all the genes present in a cell or virus
Define Genotype
Genotype is the specific genetic make-up of a cell or virus; it codes for a specific phenotype
Define Mutation
A mutation is a permanent, heritable change in the genetic material
Define Phenotype
The appearance of an organism resulting from the interaction of the genotype and the environment
How do DNA and RNA differ?
DNA is double-stranded, uses C,G,A,& T, and its sugar is deoxyribose
RNA is single-standed, uses C,G,A,& U, and its sugar is ribose
What are the 5 bases? Which are purines and which are pyrimidines?
The five bases are:
Purines-Adenine & Guanine
Pyrimidines-Thymine, Cytosine, and Uracil
What specific type of bond connects successive nucleotides in a single strand?
Phosphodiester bonds
What type of bonds connects complementary bases of two paired strands?
Hydrogen bonds
Which base pairs have two Hydrogen bonds and which have three?
Between A (adenine) and T (thymine)or U (uracil) there are 2 H-bonds, between G (guanine) and C (cytosine) there are 3 H-bonds
What is meant by the 5' and 3' directionality of the polymer?
The strand begins with the 5'-hyroxyl end and progresses towardt the 3'-hydroxyl group
Why are the strands of DNA considered antiparallel?
The strands are consdiered antiparallel because one strand is oriented 5' to 3' while the coplimentary strand is oriented 3' to 5'.
How is supercoiling maintained by Gyrase and Topoisomerase I?
Gyrase uses ATP to induce negative supercoining and Toposiomerase I relieves all supercoiling.
What are histones and histone-like proteins?
Histones (and histone-like proteins) are small, basic proteins rich in the AAs lysine and arginine that is associated with eucaryotic DNA in chromatin. They act as spools around which DNA wraps itself (to be packaged up).
What is meant by "semi-conservative" replication?
At the end of replication you are left with two double strands, each composed of one new daughter strand and one parent strand.
Why is one strand said to be "leading" and the other "lagging"?
One stand is considered to be the 'leading' strand simply because it can be extended continuously at its 3' end as the DNA unwinds. The other strand is considered the 'lagging' strand becasue it is synthesized discontinuously in the 5' to 3' direction as a series of fragments (the fragments are later joined to make a complete copy)
What is a replication fork?
A replication fork is the place at which the DNA helix is unwound and individual strands are replicated.
In replication, what is the role of DNA Polymerase I?
In replication, DNA Polymerase I digests the RNA primer and synthesizes complementary DNA to fill the gaps resulting from this RNA deletion (and proofreading)
In replication, what is the role of DNA Polymerase III?
In replication, DNA Polymerase III adds nucleotides onto the 3' end of the developing strand (moving from in the 5' to 3' direction) (and proofreading)
In replication, what is the role of RNA Primase?
In replication, RNA Primase synthesizes the RNA primer for the lagging strand
In replication, what is the role of the Primosome?
In replication,the Primosome aids the RNA primase
In replication, what is the role of DNA helicase?
In replication, DNA helicase uses ATP to undwind short stretches of helix just ahead of the replication fork
In replication, what is the role of ssDNA binding proteins?
In replication, ssDNA (single-stranded DNA) binding proteins specifically bind with DNA to keep two strands separate
In replication, what is the role of Topoisomerase I?
In replication, Topoisomerase I revieves supercoiling by changing the structure of DNA by transiently breaking 1 strand in such a way that it remains unaltered as its shape is changed. (no ATP)
In replication, what is the role of Ligase
In replication, Ligase joins the fragments of DNA produced in the lagging strand by forming a phosphodiester bond between the 3'-hydroxyl of the growing strand and the 5'-phosphate of an Okazaki fragment (use ATP or NAD+ P bond (Bac)for energy)
In replication, what is the role of Okazaki Fragments
In replication, Okazaki Fragments are formed by discontinuous synthesis of the lagging strand and are later joined to form a complete copy
What cell parameter seems to signal the initiation of replication?
How does the rate of eukaryotic replication differ from that of prokaryotes?
Eukaryotes ~ 100bp/sec
Prokaryotes ~ 1000bp/sec
Where is replication initiated?
Replication is initiated at the ori (oriC) locus (the origin of replication)
In what way do eukaryotes differ from prokaryotes in regards to where replication is initiated?
Eukaryotes have multiple ori, while prokaryotes have a single ori
What is the rolling circle mode of replication?
In the rolling circle mode of rep., one strand is nicked and the free 3'-hydroxyl end is extended by replication enzymes. As the 3' end is lengthened while the growing point rolls aroudn the circular template, the 5' end of the strand is displaced and forms an everlengthening tail, which may be converted to the dbl-stand form by complementary strand synthesis. (useful to viruses)
Is the rolling circle mode of replication bidirectional?
No, the rolling circle mode of rep. is unidirectional
What sorts of replicons (autonomously replication DNA molecules) would use the rolling circle mode of rep?
The rolling circle mode of rep. is particularly useful to viruses because it allows the rapid, continuous production of many genome copies from a single initiation event.
How do we generate induced mutations?
Induced mutations result from the application of a chemical or physical gradient
What are tautomers and how do they casue transitions?
Tautomers are sturctural isomers that are in chemical equilibrium and readily change into one another (keto, imino, enol). They cause mutation by a base taking on either the imino or enol form which changes the H bonding allowing a purine/pyrimidine to be substituted for another purine/pyrimine(Transition) or a purine to be substituted for a pyrimidine & vice versa (Transversion)
How do base analogs increase mutations?
Base analogs are sturctually similar to normal bases but they spend more time in their tautomeric form-they are incorporated into a growing polynucleotide chain-once incorporated they exhibit different base pairing properties than the base they replaced and can cause a stable mutation (Ex: 5-Bromouracil)
What types of sequences cause slipped mispairing?
Slipped mispairing generally occur where there is a short stretch of the same nucleotide. In this location, the pairing of template and new strand can be displaced by the distance of the repeated sequence leading to additions or deletions in the new strand.
What process is the cell engaged in when slipped mispairing occurs?
The cell is engaged in replication when slipped mispairing occurs.
When would slipped mispairing result in frame-shifts?
Slipped mispairing would result in frame-shifts if the repeated sequence is less than/more than 3 bases long.
How do intercalating agents such as acridine dyes cause mutations?
Intercalating agents, such as acridine dyes, distort DNA to induce single nucleotide pair insertions and deletions. These may cause a loop in DNA or frame-shift mutation.
What do I mean by DNA modifying agents?
DNA modifying agents are agents that change the existing bases in a strand of DNA. ("specific mispairing")
Define wild-type
The wild-type is the most prevelent form of a gene (normal/average)
What are forward and back mutations?
A forward mutation is a mutation from the wild-type to a mutant form. A back mutation converts the mutant nucleotide sequence back to the wild-type sequence.
How do supressor mutations differ from true reversion mutations?
A supressor mutation occurs when a mutation in a different gene overcomes a mutation and therefore regains the wild-type phenotype, while a reversion mutation is a mutation within the same gene that causes regaining of w-t phenotype
What are point mutations?
Point mutations are mutations that affect only one base pair.
What are insertions?
Insertions are mutations that occur from the addition of one or more bases
What are deletions?
Deletions are mutations that occur from the deletion of one or more bases
What are inversions?
Inversions are mutations that occur with a change in the sequence order
Why are nonsense mutations (but not missence mutations) and many insertions and deletions polar?
Polar mutations change information after/downstream of the mutation itself (ie.nonsence, frameshift, & insertions). Missence mutations only change the AA coded for by that specific codon, it doesn't change info downstream so it is not polar.
Which class of base substitution is more common?
Why may some mutations be silent?
Some mutations may be silent because the code is degenerate-many different codons may code for the same AA
How can we select for mutants?
We can select for mutants by creating growth conditions that are optimal for the mutant but will not allow growth of the parent cell.
How does replica plating work?
In replica plating, mutants are generated by treating a culture with a mutagen. The culture containing w-t and auxotrophs is plated on complete medium. After the colonies have developed, a piece of sterile velveteen is pressed on the plate surface to pick up bacteria from each colony. Then the velvet is pressed to the surface of other plates and orgs are transferred to the same position as on the master plate. After determining the location of the auxotrophic colonies growing on the replica w/complete medium, the auxotrophs can be isolated and cultured.
Is replica plating a selection or detection (screening) system?
Replica plating is a detection system.
In the Ames test, are the mutations being detected back or forward mutations?
In the Ames test the mutations being detected are back mutations.
What is meant by "biotransformation"?
“Biotransformation” means transforming a certain substance as it would be converted in a biological system. In this situation liver extract converts potential carcinogens into electrophilic derivatives that will readily react with DNA
Why might there be more mutants growing on a plate with a moderate concentration of a test substance than on a plate with a high conc of the substance?
There might be more mutants growing on a plate with a moderate concentration of a test substance that on a plate with a high concentration of the substance because a high concentration of the test substance may cause so much mutation that the organism is unable to grow up on the plate. At a moderate concentration, the test substance would cause minimal mutation (back mutation) that would allow the bacteria to grow up on a plate lacking his.
What enzymes are responsible for proofreading and when does it occur?
DNA polymerases are the enzymes responsible for proofreading; DNA polys can remove an incorrect nucleotide immediately after its addition to the growing end of the chain.
What is photoreactivation?
Photoreactivation is the repair of thymine dimmers by splitting them apart into separate thymines with the help of visible light in a photochemical reaction catalyzed by the enzyme photolyase.
What is excision repair?
Excision repair is a general repair system that corrects damage which causes distortions in the double helix. The mutation and nearby bases on either side are removed, DNA polymerase I fills in the gaps and DNA ligase joins the fragments.
What types of enzymes initiate excision repair?
AP endonucleases initiate excision repair; they recognize the location of the mutation and nick the backbone at the site.
What enzyme replaces the excised DNA?
DNA polymerase I
When is recombinational repair required?
Recombinational repair is required when gaps are left in DNA (there isn’t a good template-bad or non-existent).
Which enzyme have we discussed that participates in recombinational repair?
The enzyme RecA matches the gap with an intact strand
Which repair system is the most error prone?
SOS repair is the most error prone
What is the SOS response?
The SOS response is an induced repair system in which RecA binds to gaps in DNA and initiates strand exchange. Simultaneously RecA takes on a proteolytic function that destroys the lexA repressor protein, which derepresses many genes. Can quickly repair damage done by UV but is very error prone.
What signals initiate the SOS response?
The SOS response is initiated by the cleavage of lexA (the repressor to SOS).
Which enzyme acts as a protease in SOS?
RecA acts as a protease in SOS repair
What does LexA do?
LexA supresses SOS

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