unwinding and helicase.txt

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unwinding and helicase.txt
2012-03-26 16:36:49
helicase unwinding

unwinding and helicase
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  1. DNA topology
    • DNA organized in supercoiled loops
    • Negatively supercoiled: DNA helices wound around each other in a rihgt-handed path
    • Positively supercoiled: DNA helices wound around in a left-handed path
    • Rapid rotation of DNA helix needed as the replication fork moves
  2. Topoisomerase roles
    • Responsible for regulation of DNA pos. and neg. supercoiling
    • 1. Alter topological state of DNA
    • 2. Facilitate protein interactions w/DNA for DNA replication
    • 3. Prevent harmful excessive SUPERCOILING
  3. Topoisomerase Type 1
    • Pass one STRAND of helix through other STRAND
    • Do NOT require High energy for activation: reaction driven by energy STORED in supercoiled DNA
    • 3 subtypes
    • 1A: binds to 5' phosphate
    • 1B and 1C: bind to 3' phosphate
  4. Topoisomerase Type 2
    • Passing a region of DUPLEX DNA through another region of DUPLEX DNA
    • REQUIRES energy: Utilizes ATP to drive these conformational changes
  5. Relaxation Mechanisms
    • 1. Enzyme-bridged strand passage: proteins mediate opening of one strand to allow other thru
    • 2. Swiveling: involves rotation of DNA strands
  6. Which relaxing method do 1A, 1B and 1C use
    • Type 1A: forms transient 5' covalent intermediate and RELAXES through Enzyme-bridged strand-passage mechanism
    • Type 1A ONLY relax NEGATIVELY supercoiled DNA and require ssDNA for activity
    • Type 1B and 1C: form 3' covalent intermediates and RELAX DNA by SWIVELING
  7. Helicase
    • protein that unwinds duplex DNA by HYDROLYSIS of NTP
    • Unwinding of DNA involves: unidirectional translocation and strand separation
  8. Unwinding with helicases
    • can be catalyzed by 2 DNA helicases
    • 1. T7 gp4 helicase (unidirectional translocation 5'-3' direction)
    • 2. RepA helicase
    • these move in opposite directions along ssDNA
  9. Strand separation requirements
    • Requires breaking of hydrogen bonds b/w NUCLEOTIDE base pairs in dsDNA
    • Requires presences of NTP or ATP, AND Mg