Introduction to Nucleic Acids, DNA, Chromatin and Chromosome Structure Flashcards
Store and express genetic information and transmit it from one generation to the next
-Confer individuality
Nucleic Acids (DNA and RNA)
Joins DNA segments to form hybrid molecules
Ex: Chromosome crossing over during meiosis and the diversity of B cells
Recombination
Every cell contains the exact same DNA, but the diversity comes from the
Expression of that DNA
What nucleotide component actually carries the genetic information?
The base
How can we combat the bacteria Salmonella typhimurium?
Inactivtion of DNA adenine methylase (dam)
Blocks expression of virulent genes, prevents disease development, and induces an immune response
Inactivation of dam
Incorporated in the DNA during replication and block further DNA synthesis
-does not significantly affect the host cell metabolism
Nucleoside analogs
Hydrolyze phosphodiester bonds
Nucleases
What is the most common form of DNA?
B DNA
States that there must be equal amounts of purines and pyrimidines in the double-stranded DNA
Chargaff’s rule
Each diploid cell contains how many chromosomes?
46 (6ft of DNA)
Linear or circular B-form double helix
Relaxed conformation
Has fewer helical turns than the relaxed B-form DNA double-helix
-classified as anything less than 10 helical turns
Negative supercoil
Negative supercoils are energetically favored. The energy needed for strand separation is stored in
Supercoils
Force the eukaryotic DNA to wrap around them and generate a negative supercoil
Histones
Change the tertiary structure of DNA by transiently breaking one or both DNA strands, passing the strands through the break, and rejoining the strands
-induces “swivel points” in the DNA helix
Topoisomerases
Why do we need topoisomerases?
Positive supercoiling is too much tension, so topoisomerases come in and release the tension
How are topoisomerases able to work?
They have nuclease and ligase activity
What makes DNA gyrase (a Topo II) unusual?
It can induce negative super coils into relaxed DNA
Why is inhibition of DNA gyrase a good strategy for antibiotics?
It inhibits bacterial DNA syntheis, and since Eukaryotes do not have DNA gyrase, there are no side effects for humans
Inhibition of eukaryotic Topoisomerases is used in cancer treatment, this has signigicant side effects because it will
Lead to cell death
DNA is associated with non-histone proteins and condensed into a non-membrane bound nucleoid in
Prokaryotes
DNA is associated with histone and non-histone proteins and is condensed into a nucleoprotein complex called chromatin in
Eukaryotes
Chromatin is arranged in repeating units (like beads on a string) called
Nucleosomes
What is the structure of a Nucleosome
- ) Core: made up of DNA supercoils and histones
2. ) DNA spacer
20-80 bp of DNA between cores
-binds H1
DNA spacer
The regional compaction of chromatin is affected by
Histone modifications
As soon as DNA replication is completed, one histone H1 binds the spacer DNA which promotes
Tight packing (Solenoid)
In prokaryotes, replication is initiated by strand separation at a location rich in A:T base pairs, called the
Origin of replication
DNA unwinding is catalyzed by DNA helicase within the
Pre-priming complex
Bind DNA cooperatively to keep the single strands apart and protect them from nucleases
Single-strand DNA-binding proteins (SSBs)
What would be the affect of an inhibitor of Helicase?
Inhibited progression of DNA replication
DNA polymerase can not initiate synthesis on a totally single-stranded template. As a result, it requires an
RNA primer
Can replaced mismatched nucleotides that escaped proofreading after DNA replication
Mismatch Repair (MMR) pathway
Has 5’ to 3’ exonuclease activity, which it uses to remove RNA primers from Okazaki fragments before it fills in the gap with DNA
DNA polymerase I
ATP dependent enzyme that covalently joins Okazaki fragments
DNA ligase
Contains primase and initiates syntheses on the leading and lagging strand in eukaryotic replication
DNA polymerase α
DNA polymerase α has no
Exonuclease activity
Responsible for DNA replication on the lagging strand in eukaryotes
- associates with proliferating cell nuclear antigen (PCNA)
- has 3’ to 5’ exonuclease activity
DNA Polymerase δ
Responsible for DNA synthesis on the leading strand in eukaryotes
- associates with processivity factor of PCNA
- has 3’ to 5’ exonuclease activity
DNA Polymerase ε
Displaces the 5’ ends of primers from Okazaki fragments
DNA Polymerase δ
Consist of short non-coding G-rich DNA repeats (TTAGGG) and associated proteins
- form T loops
- Located at ends of linear chromosomes
Telomeres
Protect the ends of DNA from recognition as broken DNA and degredation
Telomeres
A ribonucleoprotein complex that adds short G-rich DNA repeats (TTAGGG) to the single stranded 3’-ends of linear chromosomes
Telomerase
Uses it’s RNA component to extend the parent strand and then its protein component (with reverse transcriptase activity) to synthesize DNA
Telomerase
Implicated in cell aging and cancer
Telomerase
In most somatic cells, telomerase is inactive. So to prevent telomere shortening and eventual chromosome end-to-end fusion, p53 induces
Cell growth arrest (stops cell from replication)
Can limit human cancer cell proliferation
Telomerae inhibitors
Inherited disease caused by reduced Telomerase activity
-affects precursor cells in highly proliferative tissues
Dyskeratosis Congenita
Patients with Dyskeratosis Congenita generally die from
Bone marrow failure
Rare inherited condition that shows accelerated telomere shortening
-Patients generally die from a myocardial infarction before age 20
Hutchinson-Gilford Progeria
Incorporation of an incorrect nucleotide or of an extra nucleotide
-None of the nucleotides are defective
Mismatch
When there is a damaged base, the Base Excision Repair (BER) pathway is activated, what does it do?
Damage base removed, sugar backbone is cut and sugar phosphate residue is removed. Gap is pilled by DNA polymerase
Repair proteins, recognize mismatches, and distinguish the newly synthesized strand with the error from the parental strand
Mismatch Repair pathway (MMR)
In prokaryotes, a mismatched DNA is recognized because it is
Not immediately methylated
How does MMR work?
Helicase and exonuclease remove the mismtched DNA and DNA poylmerase III fills the gap
90% of lynch ryndrome patients have mutations in
MSH2 or MLH1 (MMR proteins)
Reactive oxygen species (ROS), by-products of cellular metabolism and certain chemicals can cause damage to the bases in our DNA. This damage is corrected by the
Base Excision Repair pathway (BER)
Mutation that interfere with BER lead to a high risk of
Colon cancer
Premature aging disease caused by the inhibition of WRN helicase, which is involved in the BER
Werners Syndrome
Cigarette smoke contains carcinogens. Once oxidized, these compounds covalently bind to G residues in the DNA of lung cells and distort the helix. What pathway would be used to fix this?
Nucleotide excision repair pathway
The only mechanism that removes bulky DNA adduct
Nucleotide Excision repair
Used if adduct is in a transcriptionally inactive region of DNA
-Highly associated with cancer
Global Genomic NER
Used if adduct is in a transcriptionally active region of DNA
-associated with CNS disorders
Transcription-coupled NER
Xeroderma pigmentosum, a hereditary disorder resulting from defects in global genomic NER, results in extreme
Solar sensitivity and highly increased risk of skin cancer
Adducts in transcriptionally active regions of DNA block the progression of
-Halts gene transcription
RNA polymerase II
Hereditary developmental and neurological disorder associated it defects in TC-NER where there are mutations in CSA or CSB which affects recognition of stalled RNA polymerase II
Cockayne syndrome
Unlike patients with Xeroderma pigmentosum, patients with Cockayne Syndrome are not at increased risk for
Skin cancer
Repairs errors commonly caused by oxidative damage that results in a break in one strand of the DNA that is typically missing a single nucleotide
Single-strand break repair pathway
A single strand break is first recognized by
-Recruits XRCC1
PARP-1
-Poly(ADP-ribose) polymerase 1
Restores the proper 3’ OH and 5’ phosphates of the damaged DNA which enables beta-polymerase to insert the missing nucleotide
Aprataxin (APTX)
Ataxia Oculomotor Apraxia is an autosomal recessive spinocerebellar ataxia syndrome caused by mutation in the
APTX gene
Severely compromise genome stability and lead to loss of chromosome fragments in mitosis
-Cause cancer due to joining of the wrong ends
Double-stranded breaks
The major pathway to repair double-stranded breaks. Rejoins what remains of two broken DNA ends
- Does not require any sequence homology
- error prone
Non-homologous end-joining (NHEJ)
In NHEJ, frayed ends are removed (if needed) by the endonuclease activity of
DNA-PKcs:Artemis
Information on the homologous sequence is used to repair the broken DNA
-requires alignment of highly homologous DNA molecules
Homologous recombination
Results in no change in DNA sequence
-error free
Homologous recombination
Mutation in homologous recombination proteins BRCA1 or BRCA2 leads to an 80% lifetime risk of developing
Breast or Ovarian Cancer
Tumors in BRCA carriers are more sensitive to
-due to defects in DSB HR repair
Ionizing radiation
Ataxia Telangiectasia is an autosomal recessive disorder that has a propensity to develop lymphoid cancer. It is associated with a mutation in a protein that is normally activated by double stranded breaks. The protein signals the cell-cycle check point to slow the cell cycle. What is this protein?
AMT
