DNA structure and function, Replication, and Repair Flashcards
What are the components of a nucleotide.
Which are purines, pyrimidines
How many hydrogen bonds?
A base with a five carbon sugar (either 2’OH RNA, or no 2’OH, deoxyribose)
linked by an N-glycosidic bond
= nucleoside
The addition of the phosphate group to the sugar via a phosphoester bond. makes it a nucleotide.
Purines; AG
Pyrimidine: CT
3 hydrogen bonds: CG
2 hydrogen bonds: AT
Salmonella causes food poisoning. What was a conceptual treatment?
hint, methyl groups induce expression on DNA for viruses
some bases are methylated, and this allows protein to recognize the DNA for expression.
Inactivation of DNA adenine methylase blocks the expression,
Gut colonizing pathogens all carry methylases, meningitis, cholera…etc
Acyclovir - what is it a type of?
what is it used for?
It is a nucleoside analog that doesn’t allow for elongation, missing 3’ OH. You let the organism incorporate it into a nucleotide.
Used for Herpes Simplex virus infections.
Analog of deoxyguanosine
AZT what is it used for? Dallas buyers club.
analog of thymidine
Used to treat HIV and AIDS, like acyclovir. It blocks further DNA synthesis.
Does not significantly affect host cell metabolism which is good and odd compared to the movie…
What is the most common form of DNA?
How many bases per helical turn?
How many angstroms is each turn?
B form
10 bases per helical turn
34 angstroms each turn.
Certain anti-cancer drug interferes with DNA and RNA synthesis by intercalating what?
the minor groove.
What are the benefits to negative supercoils?
How are they formed?
Easier strand separation, energetically favorable, and the energy for strand separation is stored in the super coil.
Formed by partial unwinding of the double helix. Then restoring complementary basepairing. End result is one less helical turn. 100 basepairs and 10 helical turns. Remove a helical turn, 9 helical turns, the Molecule folds on itself.
When eukaryotic histone proteins bind, they force the DNA to wrap around them and this generates a negative supercoil.
What are DNA topoisomerases and generally what do you need to know?
They introduce swivel points into the DNA. Topo 1 only does single strand breaks and strings one strand through the other. Removes positive and negative supercoiling.
Topo 2 is a double strand break, and does both as well. REQUIRES ATP AND TOPO 1 DOESN’T.
Gyrase is exclusive to prokaryotes and it can pump negative supercoils.
How are topoisomerase inhibitors used in anti-bacterial therapy and anti-cancer therapy?
inhibits bacterial DNA synthesis by blocking gyrase, the tension becomes too much for strand separation. Eukaryotes don’t have gyrase so no side effects.
anticancer, remove the nick sealing portion of the topoisomerases 1 and 2, so it becomes DNA breaking agents! But this leads to cell death in eukaryotic cells as well
Describe the structure of histones. Include linker.
What is the order of compaction?
An octomer, two of each. H2A, H2B, H3, H4, they’re lysine rich.
There is a DNA spacer, 20-80 bp of DNA between histones and it binds H1. This linker is allows the cores to become closer and coil.
nucleosomes = 10 nm fiber > 30 nm fiber (solenoid) = then DNA loops coiled around a protein scaffold.
Describe the mechanism of prokaryote replication.
The replisome, carries out replication. Strand separation begins at the origin of replication (usually AT rich), DnaA recognizes the AT region and uses ATP to locally melt it. This forms two replication forks, helicase binds near the forks and uses ATP to force the strands open. Single strand binding proteins stabilize the single strands to keep them apart and protect them from degradation.
Topoisomerase 2 works ahead of the fork to relieve tension .
Which polymerase catalyzes chain elongation in prokaryotes?
Which exonuclease activity does it contain.
What does DNA pol 1 do?
DNA polymerase III catalyzes 3’-5’ bonds with OH attack 5’ phosphate.
3’ to 5’ exonuclease activity. DNA is made 5’ to 3’ so this lets it rewind.
DNA polymerase 1 has 5’ to 3’ exonuclease activity so it removes the primers ahead of pol 3. (It also has 3’ to 5’ exonuclease activity and can synthesize DNA)
Which molecules are ATP dependent?
Topoisomerase 2’s
Helicase
DNA ligase.
What is different about eukaryote replication? Especially polymerases.
Multiple origins of replication,
DNA is associated to histones,
Has telomeres
Eukaryotic DNA polymerases,
Pol alpha (contains primase, initiates synthesis on leading and lagging strands)
Pol delta - elongates lagging strand (has 3’ to 5’ exonuclease activity)
-displaces the primer on lagging strand , the flap that forms is degraded by exonucleases (substitutes function of Pol 1 in prokaryotes)
Pol epsilon (elongates leading strand, same exonuclease activity) if dysfunctional pol delta CAN SUBSTITUTE.
What happens in nucleosomes as fork advances?
Histones will remain loosely associated to a parental strand, new histones are synthesized simultaneously with DNA replication and nucleosomes reform behind the replication fork
Describe telomeres. What do t-loops do?
They are at the end of linear chromosomes.
Telomerase contains an RNA code and reverse transcribes to extend to 3’ ends. The primer will be inserted on the 3’ end to synthesize from 5’ to 3’. Once replication is over, the primer is removed and there is a 3’ overhang which will get digested. This is so only nonsense DNA is lost.
Telomeres are short noncoding G-rich sequences (TTAGGG) and they form t-loops.
Protect the ends from degradation, prevents recombination, and end to end fusion.
Describe the role of telomeres in aging and cancer
telomerase is active in all cells before birth and remain active in stem cells and germ cells.
When telomeres get too short that end to end fusion can occur, DNA damage sensors arrest cells (p53), and serves as a mitotic clock by monitoring telomere length.
Telomerase is usually active and p53 is deactivated in cancers. Telomerase inhibitors.
What are the effects of Dyskeratosis congenita (reduced telomerase activity)
affects precursor cells in proliferative tissues :
Hair = baldness,
Intestinal crypt = gut disorders
Basal layer of epidermis = abnormal skin pigmentation.
Bone marrow progenitor cells. (fail to produce new blood cells so patients usually die from bone marrow failure.
Hutchinson-Gilford progeria
cause and effect
accelerated telomere shortening,
aged skin, alopecia, short stature, early death by MI.
What kind of repair mechanism is best for each of the following?
damage to a single base? replication errors bulky adducts oxidative damage - single strand break ionizing radiation, oxidizing agents.
damage to a single base = BER
replication errors = MMR
bulky adduct = GG NER, TC NER
single strand break repair
doulble strand break = homologous, non homologous.
Describe MMR
mismatch, no nucleotide damage.
MSH2/6 recognizes mismatch while MSH 2/3 recognizes insertions and deletions. They will recruit MLH-1/PMS2 which are endonucleases.
Prokaryotes recognize parental strand by methylation and eukaryotes perhaps recognize the nicks on lagging strand.
The nucleases nick on either side of the mismatch. The helicase unwinds so it is singlestrand and exonuclease digests it. DNA pol III and ligase fill the gap.
Defects in MMR
Lynch syndrome: significantly increased risk of colorectal, endometrial, ovarian, skin, gastric, renal cancers.
BER
ROS (reactive oxygen specifies can damage our bases) example cytosine deaminated into uracil.
First the damaged base is recognized by base specific glycosylase, uracil DNA glycosylase. Which cleaves the N-glycosidic bond and removes the base.
AP endonuclease cleaves the sugar phosphate backbone.
Deoxyribose phosphate lyase removes the residue. DNA pol I fills the gap
Defects in BER
mutation in DNA glycosylase leads to very high risk for colon cancer - adenomatous colorectal polyposis syndrome.
Werners syndrome: for some fucking reason helicase isn’t mentioned for BER, however WRN helicase is necessary for BER…
autosomal recessive disorder ft. premature aging, skin atrophy, hair loss and cancer disposition.
Wrn is also involved in processing telomeric DNA, activating DNA damage responses.
