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In vivo DNA binding pattern of the Polycomb Txn Factor

In vivo DNA binding pattern of the Polycomb Txn Factor. What are the genes to which it binds? How does it affect these genes?. 3. What determines where it Binds??. Nitrogenous base. Sugar. Phosphate. 1. Evidence for the Double Helix. helical. 1. Fiber Diffraction data :

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In vivo DNA binding pattern of the Polycomb Txn Factor

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  1. In vivo DNA binding pattern of the Polycomb Txn Factor • What are the genes to • which it binds? • How does it affect these • genes? 3. What determines where it Binds?? 免疫学信息网 http://www.immuneweb.com

  2. Nitrogenous base Sugar Phosphate 1 免疫学信息网 http://www.immuneweb.com

  3. Evidence for the Double Helix helical 1. Fiber Diffraction data: -Helical geometry -3.4 A º spacing (1Aº = 10-10 m) -34 A º pitch 10 layer Lines Between Cross Patterns (10 Residues Per turn) 2. Structure of dCTP 3. Base Tautomerism • Chargaff rules • - A=T, G=C 1A 免疫学信息网 http://www.immuneweb.com

  4. NIH (not in handout) 免疫学信息网 http://www.immuneweb.com

  5. -2’-deoxyribose 2 免疫学信息网 http://www.immuneweb.com

  6. Sugar “Pucker” Conformations A DNA B DNA 3 免疫学信息网 http://www.immuneweb.com

  7. Pyrimidines Purines 4 免疫学信息网 http://www.immuneweb.com

  8. Base Tautomerization G (Keto) G (Enol) A 99.99% 0.01% 5 免疫学信息网 http://www.immuneweb.com

  9. 9 1’ Base Adenine Guanine Thymine Cytosine Nucleoside (Deoxy)adenosine (Deoxy)guanosine (Deoxy)thymidine (Deoxy)cytidine Nucleotide (d)A (mono, di-, tri) phosphate (d)G (mono, di-, tri) phosphate (d)T (mono, di-, tri) phosphate (d)C (mono, di-, tri) phosphate 6 免疫学信息网 http://www.immuneweb.com

  10. A very useful number:660 免疫学信息网 http://www.immuneweb.com

  11. Rotation About the N-Glycosidic Bond N3 A A,B DNA Z DNA (G only) 7 免疫学信息网 http://www.immuneweb.com

  12. 8 Phosphodiester Backbone 免疫学信息网 http://www.immuneweb.com

  13. Rise 3.4 Å Minor Groove B-DNA: A right Handed double helix Why? Major Groove Pitch 34 Å 10.4 bp/turn 9 免疫学信息网 http://www.immuneweb.com Width 20 Å

  14. Twist 36° 9 免疫学信息网 http://www.immuneweb.com

  15. Major Groove 8.5 Å 11.7 Å Minor Groove 7.5 Å 5.7 Å 10 免疫学信息网 http://www.immuneweb.com

  16. 11 免疫学信息网 http://www.immuneweb.com

  17. Note to self:Discuss forces that affect helix formation 免疫学信息网 http://www.immuneweb.com

  18. C-G T-A NIH 免疫学信息网 http://www.immuneweb.com

  19. Pitch Handedness Base Inclination A B Z 12 免疫学信息网 http://www.immuneweb.com

  20. B DNA A DNA Major Minor Major Minor dx = 0.8 Å dx = -4 Å Base Displacement Determines Groove Depth Z DNA Major Minor dx = +3-4 Å 13 免疫学信息网 http://www.immuneweb.com

  21. A B Z Mi Ma Ma Mi Mi Ma 12 免疫学信息网 http://www.immuneweb.com

  22. Z-DNA Phosphate Backbone is Kinked A B Z 14 免疫学信息网 http://www.immuneweb.com

  23. 15 免疫学信息网 http://www.immuneweb.com

  24. Question: is all B-DNA structurally identical? Implications of structural variation Implications of flexibility 免疫学信息网 http://www.immuneweb.com

  25. Degrees of freedom: 7 Torsion angles and sugar conformation (Rigid) 5’ 3’ 16 免疫学信息网 http://www.immuneweb.com

  26. Structural Variation Defined by Bases normal frequent never Common Common Never (except in intercalation) 17 免疫学信息网 http://www.immuneweb.com

  27. Propeller Twist Maximizes Base Stacking 5’ 3’ 5’ 3’ 3’ 5’ 5’ 3’ NIH 免疫学信息网 http://www.immuneweb.com

  28. Buckle Propeller Twist 18 Textbook Real Life 免疫学信息网 http://www.immuneweb.com

  29. Naturally Occurring Variations in Roll, Slide, Twist 19 免疫学信息网 http://www.immuneweb.com

  30. 5’ 5’ C T A G 3’ 3’ Pyrimidine-Purine Steps Have Little Base Stacking Step Definition: Going along one strand of DNA in 5’to 3’ direction Four Possibles: P-Y, P-P, Y-P, Y-Y 19A 免疫学信息网 http://www.immuneweb.com

  31. 5’ 3’ A G C 5’ T 3’ Purine-Pyrimidine Steps Have Extensive Base Stacking 19B 免疫学信息网 http://www.immuneweb.com

  32. For further reading on effects of sequence on structure, “Understanding DNA-The Molecule and How it Works” By Calladine and Drew Major Conclusion: DNA structure can depend on sequence In predictable, yet complicated ways. Therefore, DNA binding proteins can recognize structure, And they can be designed to bind to highly flexible DNA. 免疫学信息网 http://www.immuneweb.com

  33. DNA Topology**Johannes’ Favorite Subject(Students’ least favorite subject) 免疫学信息网 http://www.immuneweb.com

  34. DNA Unwinding Causes Topological Problems Unwound Parental Duplex (Transcription) Over- Wound region 20 免疫学信息网 http://www.immuneweb.com

  35. More Topological Problems 21 免疫学信息网 http://www.immuneweb.com

  36. Properties of Topoisomerases 22 免疫学信息网 http://www.immuneweb.com

  37. Strand Passage Model for Topo I Covalent Tyrosine-5’P Strand Passage Re- ligation Unwound Complex Cleavage Complex L=2 L=3 23 免疫学信息网 http://www.immuneweb.com

  38. Topo I Reactions 24 免疫学信息网 http://www.immuneweb.com

  39. Model for Topo II Mechanism 25 免疫学信息网 http://www.immuneweb.com

  40. Topo II Reactions 26 免疫学信息网 http://www.immuneweb.com

  41. For a good treatment of topos, see the book: “DNA replication” Arthur Kornberg and Tania Baker 免疫学信息网 http://www.immuneweb.com

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