Chapter 4.5/4.6

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mse263
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108041
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Chapter 4.5/4.6
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2011-10-11 23:33:38
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Lecture
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MCBI Exam 1
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  1. DNA helicase
    motor proteins that move directionally along a nucleic acid phosphodiester backbone, separating two annealed DNA strands using energy from ATP hydrolysis; start the process at ORIs (origins of replication) who's nucleotide sequence is often A/T rich
  2. Primase
  3. DNA polymerase
    enzyme that catalyzes polymerization of deoxyribonucleotides into a DNA strand; unlike RNA polymerase they can't initiate chain synthesis de novo and require a short preexisting RNA/DNA strand called a primer; are best known for their feedback role in DNA replication, in which the polymerase "reads" an intact DNA strand as a template and uses it to synthesize the new strand; this process copies a piece of DNA; newly polymerized molecule is complementary to the template strand and identical to the template's original partner strand
  4. DNA ligase
  5. conservative mechanism
    by this mechanism two daughter strands would form a new double-stranded DNA molecule and the parental DNA duplex would remain intact (NOT the way DNA replicates)
  6. semiconservative mechanism
    parental strands are permanently separated and each forms a duplex molecule with the daughter strand base-paired to it; HOW DNA is REPLICATED
  7. leading strand
    because DNA polymerase can only add dNTP's in the 5' to 3' direction, only synthesis of the leading strand (which moves in the 5' to 3' direction) is continuous
  8. lagging strand
    replication of the template DNA strand to form the lagging strand must happen in the opposite direction in which the replication fork is moving; the cell accomplishes this by synthesizing a new primer every few hundred bases; said discontinuous segments = Okazaki fragments; the RNA primer of each fragment is removed as replaced by DNA chain growth from neighboring fragment and they are joined by DNA ligase
  9. SV40 DNA
    • the genome of a small virus that infects monkeys; used to study the proteins that participate in DNA replication; the molecular machinery that replicates SV40 only uses one viral protein (large T-antigen: and the REST are from the host cell's DNA!
    • -L. t-antigen = hexamer, meaning it has 6 subunits
  10. DNA polymerase α
    part of a complex along with primase; together they make the short RNA primers necessary for replication; Pol α is directly responsible for elongation the primer with deoxynucleotides (results in mixed RNA/DNA primer)
  11. DNA polymerase δ (delta)
    MAIN polymerase used for extending/adding nucleotides to the daughter strand being created in DNA replication; forms a complex with Rfc (replication factor C) and PCNA (proliferating cell nuclear antigen) that DISPLACES Pol α after primer synthesis
  12. PCNA
    proliferating cell nuclear antigen; homotrimeric (composed of three identical units of polypeptide) protein that has a central hole through which the DNA passes; preventing the PCNA-Rfc-Pol δ complex from dissociating from the template (and replication from abruptly ending)
  13. RPA
    replication protein A; protein that binds single-stranded DNA (ssDNA) preventing it from winding back on itself or forming secondary structures so DNA polymerase can continue to replicate it; Pol α and Pol δ can dislodge them as they synthesize complementary DNA strands
  14. ribonucleases H and FEN
    remove the ribonucleotides at the 5' ends of Okazaki fragments; removed bases are replaced by DNA pol δ as it extends the upstream ('backwards') fragments; ligase couples the still yet to be attached fragments
  15. telomerase
    reverse transcriptase that adds DNA sequence repeats ("TTAGGG" in vertebrates) to 3' end of template DNA strands in the telomere (end) regions; it carries its own RNA molecule, which it uses as a template when it elongates telomeres on the parental strand; after elongation, DNA pol α with the help of a primer attaches corresponding nucleotides to the lagging strand (in the same way okazaki fragments are synthesized); this region = non-coding DNA and prevents loss of important DNA from chromosome ends
  16. when you start with two large t-antigen hexameric helicases and they both move away from the origin, there is:
    • leading strand AND lagging strand synthesis at both forks:
  17. now starts 4.6
    :)
  18. recombination
    a mechanism for repairing double-stranded DNA breaks (can also be used as a way to generate new combinations of maternal and paternal genes)
  19. defects in DNA repair mechanisms and ______ are closely related
    cancer; when repair mechanisms don't function, mutations accumulate in the cell's DNA; if these mutations affects genes that are needed for regulation of cell division, cells can begin to divide uncontrollably
  20. DNA polymerase prevents mutations:
    if DNA pol accidentally adds an incorrect nucleotide at the 3' end of the daughter strand, base pairing doesn't occur between template and daughter strand; this lack of hydrogen bond formation causes the polymerase to stop adding nucleotides and instead the 3' end of the new strand is positioned at the DNA pol's EXONUCLEASE site: the mispaired base is removed and the 3' end is transferred back to the DNA pol site and the region is copied CORRECTLY
  21. number of DNA damage events in a single human cell per day:
    • ranges from 104 to 106
    • WOAH!
  22. point mutations
    cause spontaneous mutations; involve a change in a single base pair in the DNA sequence
  23. frequent point mutation: the demamination of Cytosine to Uracil
    • -in humans the most common is actually the conversion of 5-methyl C to T
    • -in this system, the repair system (a DNA glycosylase) has evolved to recognize that a G * T most often means the point mutation is localized to the T base
    • -glycosylase flips the T our of the helix, hydrolyzes the bond, and then an AP (apurinic) endonuclease cuts the DNA at this site
    • -a specialzed DNA polymerase finishes the job by removing the deoxyribose phosphate (without base) and replacing it with C base
  24. ionizing radiation (2 types) v. UV radiation
    x-rays and gamma rays cause DOUBLE-strand breaks in DNA

    UV radiation from sunlight causes distortions in the DNA double helix that interfere with replication/translation
  25. the main conceptual problem with any (base, mismatch, nucleotide) excision repair is...
    which is the NORMAL and which is the mutant DNA strand
  26. DNA glycosylase
    catalyze the first step of base excision repair, the mechanism by which damaged bases in DNA are removed/replaced; specifically, they remove the damaged nitrogenous base by flipping it out of the double helix while leaving the sugar-phosphate backbone intact, creating an apurinic/apyrimidinic (AP) site; last step = cleavage of the N-glycosidic bond
  27. YOU NEVER FINISHED MAKING FLASH CARDS

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