DNAI

Degrees of freedom: 7 Torsion angles and sugar conformation
(Rigid)
5’
3’
16
Structural Variation Defined by Bases
normal frequent never
Never (except in intercalation)
DNA Topology*
*Johannes’ Favorite Subject (Students’ least favorite subject)
DNA Unwinding Causes Topological Problems
Unwound Parental Duplex
(Transcription)
5’ C
T
G
5’
3’
3’Baidu Nhomakorabea
A
19A
Purine-Pyrimidine Steps Have Extensive Base Stacking
3’
5’ A C
3’ G T
5’
19B
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.
-2’-deoxyribose
2
Sugar “Pucker” Conformations
A DNA
B DNA
3
Pyrimidines
Purines
4
Base Tautomerization
G (Keto)
G (Enol)
A
99.99%
0.01%
5
9
1’
Base Adenine Guanine Thymine Cytosine
dx = +3-4 Å
13
A
B
Z
Mi
Ma Mi
Ma
Mi
Ma
12
Z-DNA Phosphate Backbone is Kinked
A
B
Z
14
15
Question: is all B-DNA structurally identical?
Implications of structural variation Implications of flexibility
C-G
T-A
NIH
A
Pitch
B
Z
Base Inclination
Handedness
12
Base Displacement Determines Groove Depth A DNA B DNA Z DNA
Major
Minor
Major
Minor
Major
Minor
dx = -4 Å
dx = 0.8 Å
2. Structure of dCTP 3. Base Tautomerism 3. Chargaff rules - A=T, G=C helical
10 layer Lines Between Cross Patterns (10 Residues Per turn)
1A
NIH
(not in handout)
Common
Common
17
Propeller Twist Maximizes Base Stacking
5’ 3’
5’ 3’
3’ 5’
3’ 5’
NIH
Buckle
Propeller Twist
Textbook
Real Life
18
Naturally Occurring Variations in Roll, Slide, Twist
Pitch 34 Å 10.4 bp/turn
Minor Groove Major Groove
Width 20 Å
9
Twist 36°
9
8.5 Å
11.7 Å
Major Groove Minor Groove
7.5 Å
5.7 Å
10
11
Note to self: Discuss forces that affect helix formation
19
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
6
A very useful number: 660
Rotation About the N-Glycosidic Bond
N3
A
A,B DNA
Z DNA (G only)
7
8
Phosphodiester Backbone
Rise 3.4 Å
B-DNA: A right Handed double helix Why?
Announcements
Assigned Papers: download from website: http://tfiib.med.harvard.edu/bcmp200/ Sign-up sheets
Facts (“vocabulary”) Concepts Techniques Quantitation
OverWound region
20
More Topological Problems
21
Properties of Topoisomerases
22
Strand Passage Model for Topo I
Covalent Tyrosine-5’P
Unwound Complex
Cleavage Complex
Nitrogenous base Sugar Phosphate
1
Evidence for the Double Helix
1. Fiber Diffraction data: -Helical geometry -3.4 A º spacing (1Aº = 10-10 m) -34 A º pitch
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
Strand Passage
Religation
L=2
23
L=3
Topo I Reactions
24
Model for Topo II Mechanism
25
Topo II Reactions
26
For a good treatment of topos, see the book: “DNA replication” Arthur Kornberg and Tania Baker
In vivo DNA binding pattern of the Polycomb Txn Factor
1. What are the genes to which it binds?
2. How does it affect these genes?
3. What determines where it Binds??