Biochem LEC 4-6

  1. How does replication termination differ for linear vs. circular chromosomes
    • linear- replication ends when the the polymerase reaches the telomeres
    • circular-ter sites are utilized by TuS and Tus inhibits helicase
  2. function of telomeres
    • protect the ends of the chromosome from deterioration 
    • prevents fusion with neighboring chromosomes
  3. how are telomeres synthesized
    • telomerase expressed in embryonic, stem, and cancer cells
    • not active in adult tissue
  4. DNA damage vs. mutation
    physical abnormality in the DNA (ex. single/double strand breaks)

    mutation is a change in the base sequence of the DNA
  5. hereditary diseases related to defects in DNA repair systems
    • colon cancers have defects in NER repair enzymes
    • Defects in NER can also lead to xeroderma pigmentosum(skin cells unable to repair UV-damaged DNA)
    • Rad51 (eukaryotic homolog of Rec A) interacts with Brca 1 and 2
  6. 3 General Post-replication DNA repair strategies
    • Direct Repair
    • Nucleotide Excision Repair (NER)
    • Recombination repair
  7. Direct repair
    repairs modified bases without their removal
  8. Example of Direct repair
    • Pyrimidine dimers(cyclobutyl rings)formed by UV light
    • Inhibits replication and transcription
    • DNA photolysases bind and absorb a UV photo, transfer e- to break dimer
  9. NER
    Nucleotine Excision Repair
  10. Example of NER
    • mismatch in new DNA strand
    • various systems excise the region by nicking and excising 
    • Pol III and ligase repair 
    • Dam methylase is used to methylate new correct strand
  11. Recombinational repair
    allows damaged chromosome to be repaired using a sister chromatid or homologous chromosome as a template
  12. Main enzyme in Recombinational repair and role and properties
    • RecA
    • protein-mediated DNA synapsis
    • has multiple DNA binding sites to hold a single strand and double helix together
    • DNA-depedent ATPase
  13. what is recombinant DNA
    DNA that has been formed artificially by combining constituents from different organisms
  14. Limitations of plasmid
    generally accept  <10kb of extra DNA
  15. main differences between bacterial plasmids and YAC's
    • Yacs: used for cloning large DNA segments
    • contains ARS (eukary oris), TEL, and CEN

    Plasmids contains oris, but no CEN and TEL
  16. uses for cloned DNA
    • helps detail information about entire genomes
    • new avenues for identifying and isolating genes and proteins
    • helps understand the function of newly discovered proteins
    • helps discover new biochemical processes
    • new approaches to understand metabolism
  17. how is reverse transcriptase used for cloning DNA
    you can receive an mRNA and use RT to obtain DNA fragment
  18. RNA synthesis
    • RNA polymerase synthesized from 5' to 3' on the antisense strand
    • no primer required
  19. effects of transcription on DNA topology
    • forms knot-dna is unwound at the leading edge of the transcription bubble and rewound at its rear
    • topo I- relaxes negative super coils in front 
    • dna gyrase- relaxes positive supercoils in rear
  20. difference  and similarity between eukaryotic and bacteria rna polymerases
    • eukary: have separate polis for distinct RNA classes
    • bacteria: only one RNAP

    both have a very similar claw like structure
  21. Eukaryotic RNA polymerases and properties
    • POL I- makes rRNA, no sensitivity to amantin
    • POL II-mRNA, high sensitivity
    • POL III- tRNA and snRNA, moderate sensitivity
  22. Steps of transcription
    • binding of RNAP
    • initiation of polymerization
    • chain elongation
    • chain termination
  23. Steps in Transcription Initiation step
    • migration and promoter recognition 
    • formation of closed complex
    • unwinding of DNA to form open complex
    • initiation of RNA synthesis
    • σ-subunit is released and RNA is elongated
  24. First nucleotide that attaches to RNAP to initiate synthesis
    the pppGpN-OH, purine triphosphate
  25. how does deviation from -10 and -35 nt affect promoter activity
    reduces promoter utilization
  26. the two mechanisms of transcription termination
    • rho factor-dependent
    • rho factor-independent
  27. differentes between two transcription termination
    • rho dependent- uses rho termination factor (ATP-dependent helicase)
    • Rho independent- termination sequence forms a palindrome due to run of weak A::U bonds 
    • allows RNA to fall off DNA
Card Set
Biochem LEC 4-6