A vs B vs Z DNA, Triplexesand Quadruplexes

N3

NN1

N

O N

O6N2

H

H

HN

N9

O

O

O

P

OO

O

P5'O O

O--O

H

5'

3'

H

3'

GO

OC

O

O

P

5'

P

5'

3'

3'

O

O-O

-O

5'

3'

G

5'

3'

C

O

O

O

P

N9 N3 HN2

3'

5'

OO-

O

O

N1O6

O

P

3'

5'

O O-

DIFFERENT PERSPECTIVES ON THE WATSON CRICK BASE PAIR (BP)

Looking into minor groove

minor groove side

MAJOR GROOVE SIDE

C2 pseudo axis of symmetry

TOP VIEW

SIDE VIEW

SCHEMATIC VIEW

5'-G-3'

3'-C-5'

Figure: wcbp1.cw2

5' 3'G

5'3'C

Predicting Strand Orientation From Base Pair Geometry and Glycosyl Angle

Helical parameters - [Figure 2-14]Helix axis. Line at the center of the helix parallel to thedirection of the helix.

Pitch - The distance traveled along the helix axis for a completeturn.

Helical repeat - The number of monomer units per completeturn.

Twist angle - Angle at which the pseudo dyad axis rotates ongoing from one base pair to the following base pair.

Diameter - Twice the largest radius in the helix.

Roll angle - Deviation of the angle about C6-C8 for a base pairthat is assigned zero for that in which the base pair plane isperpendicular to the helix axis.

Tilt - Deviation of the pseudodyad axis from 0 for that in whichthe base pair plane is perpendicular to the helix axis

Propellar twist - Angle between the planes of each individualbase of the base pair.

Displacement D - The distance between the helix axis and thecenter of the base pair.

Center of the base pair - The point where the C6-C8 bondcrosses the pseudodyad axis.

Helical rise per residue, h - Distance along the helix axisbetween the residues.

Base Pair Angle Definitions

Figure angledef.ppt

Dickerson Dodecamer (bdl001.pdb)

Figure BDNA1.ppt

Arnott A RNA Structure (arn0035.pdb)

Figure Adna.ppt

Comparison of groove width and depth

MAJOR

minor

MAJOR

MAJORMAJOR

minor

minor

A DNA B DNA

Figure AvsB.ppt

O

OP

O P

Base

O

OP

O P

Base

C2'-endo, 2E C3'-endo, 3E7.0 A 5.9 A

B DNA A DNA

3.4 A h = 3 A

Orientation of base pairsrelative to helix axis

h = 3.4 A3.4 A

file: c2ec3e1.cw2

Relationship between sugar pucker and helix type

C2’-endo C3’-endo

Contrasting A and B Helices A helix B helix helical repeat 11 10.5 displacement, d 4.4 -0.2 - -1.8 helical rise/residue, h 2.6-3.3 3.4 tilt 10-20 -6 twist 30-33 36-45 propellar twist sugar pucker C3’-endo, 3E C2’-endo and others, 2E intra chain P-P distance

5.9 7.0

Major groove width 2.7 11.7 depth 13.5 8.5 minor groove width 11 5.7 depth 2.8 7.5

Table: AvsB.doc

h

helic

al re

peat

minor grooveclash at 5'-Py-Pu-3'

major grooveclash at 5'-Pu-Py-3'

minimizingminor grooveclash at 5'-Py-Pu-3'

Figure calrule1.ppt

Consequences of propeller twisting on local B DNA structure and how DNA responds

Shifting BPover to minimizeclashes

Clash due to largetwist angle

Clash minimizedby decreasingtwist angle

Figure calrule2.ppt

Sequence-Dependent Variation of DNAStructure: Callidine’s rules

Local DNA structure can be viewed as aresult of two opposing effects:

1. The drive to optimize H-bonding and base stackinginteractions.

2. The drive to minimize unfavorable VDW interactionsbetween purines that result from the positive propellartwisting of the base pairs at Pu-Py (RY) and Py-Pu (YR)sequences.

DNA compensates for the clashes by bothlocal and coupled responses:1. Local Responses

a. Decrease propellar twist to 0b. Slide base pair over (reduce d)

2. Coupled Responsesa. Reduce relative twistb. Reduce relative roll angle

Document: calladine.doc

Major Groove

Minor Groove

local responses (at a base pair)

R Y Y R

propellar twist (always decrease)

-1 -1 -2 -2

d (open purine d, decrease pyrimidine d)

+1 -1 -2 +2

coupled responses (between base pairs)

x R Y x x Y R x

twist angle (always reduce) +1-2 +1 +2 -4 +2

roll angle (+ value defined as opening in the major groove; changing the central roll angle must be equally compensated by the adjacent roll angles)

+1 -2 +1 -2 +4 -2

table calladine.ppt

Relative magnitude of responses

CD of poly (dG-dC) at pH 7 25 oCSolid line: 0.2 M NaClDotted line: after addition of more NaCl

240 260 280 300wavelength (nm)

e L-e

R

-5

+3

Fig. 4-0

Split CD curve explained by exciton chirality rule

Rich hexamer Z DNA structure (zdf002.pdb)

Figure: zdna1.pptFigure 4-1

Rich hexamer Z DNA structure (zdf002.pdb)

Figure: zdna2.ppt

Minor Groove

Major Groove

Figure 4-2

Rich hexamer Z DNA structure (zdf002.pdb)

Figure: zdna3.ppt

CpG stepC3G4-C9G10

GpC stepG4C5-G8C9

anti

syn

Interstrandbase stacking

Intrastrandbase stacking

Figure 4-3

Structural Features of Z DNA

1. No major groove, just a surface 13.8 A wide, 2 A deep.

2. Very deep and narrow minor groove, 3.7 A wide, 8.8 Adeep.

3. 18 A wide helix (19 A for B, 23 for A)

4. Interchain phosphate-phosphate distance of 7.7 A; highcharge density.

5. 12 bp/turn, 7.4 A/dinucleotide repeat

6. Watson Crick base pairing between C and G.

7. Dinucleotide repeat with an intrastrand GC pi stack at 5’-GC-3’ steps and an interstand CC pi stack at 5’-CG-3’steps.

8. Left-handed stacking at 5’-GC-3’ site with at twist of -50o and -15o twist at 5’-CG-3’ steps.

9. Helix axis is dislocated at 5’-CG-3’ steps.

10. G is in a syn glycosyl conformation, C is in an anticonformation.

Figure: zfeature.cwg

NN

N

N

O

N

H

HH

O

O

O

N O

O

O

N

O

NHH

file: b2z1.cw2

3'-up

5'-down5'-up

3'-down

NN

N

N

O

N

H

HH

O O

O N OO

O

N

O

NH

H

3'-down5'3'-up

Base pairflips by 180o

anti anti

5'-up

OC

O

O

5'

3'

OG

O

O

OG

O

O

OC

O

O5'

3'glycosylbond rotates

wholenucleosiderotates

SIDE VIEW

TOP VIEW

Flipping of Base Pairs in B to Z transition

Figure 4-4

GC GC

GC GC

GC GC

GC GC

CG CG

CG CG

CG CG

CG CG

3' 5'

5' 3'

3'

5'

5'

3'

GC

GC

GC

GC

CG

CG

CG

CG

B DNA Z DNA

Figure: b2z1.cdr

=O

aa

a

aa

aa

aa

a

a a

a a

a a

a a

a a

a a

a a

s

s

ss

ss

ss

a= anti glycosyls = syn glycosyl

Conformational Transition BetweenB and Z DNA

Figure 4-5

A nanomechanical device based on B to Z transtion

Donor and acceptor molecules (fluorescein and Cy3) are attached to a DNA molecule containing a (GC)20section. When in B form the two dyes are close and show strong FRET, when in Z form, the DNA unwinds by about 3.5 turns, and extends about 6 A, changing the distance by 20-60 A, and greatly lowers the FRET.

Nature. 1999, 144-6

NN

NN

O

NHH

H

OO

OP

OO

O

O-

C

OPOO

HOMg

O

HH

H2O

OH2

OH2

H2OH

O

Figure: mg_zdna1.cw2

Ion (mM) poly d(G-C) poly d(G-m5C)

Na+ 2500 700

Mg2+ 700 0.6

Ca2+ 100 0.6

Ba2+ 40 0.6

Co(NH3)63+ 0.02 0.005

EtOH 60% v/v 20%

Mg2+ + 10% EtOH 4 mM -

Mg2+ + 20% EtOH 0.4 mM -

Minimum Salt ConcentrationRequired to Form Z DNA

Figure 4-6

NN

N

N

O

N

H

HH

H

dR

CH3

CH3

dR

H

HH

O

N

NN

m5Cm7G

Figure: zfactors.cw2

Structural factors favoring Z DNA formation

electrostatics(Z DNA has highercharge density)

hydrophobicity(methyl group occupieshydrophobic pocket)

less severe steric interactionsin the syn conformation whichis the conformation at the purinesite in Z DNA

N

NN

N

O

N

HH

HN

RO

RO O OCH3

bad steric interactionsin the anti conformation

N

NN

N

O

N

H

H

H

N

ORO

RO

CH3

O

Figure 4-7

Figure 4-8

NN

NN

N

N

O

N

H

HH

H N

O

dR

HH

H

H

dR

NN

NN

N

N

O

H

H N

O

dR

HH

H

H

dR

NN

NN

N

N

N

N

H

HH

H O

O

dR

H

H

HH

dR

inosine

2-aminopurine

Figure: 2ap_ino1.cw2

Polymer C2-NH2 group Helix

d(A)•d(T) no B

d(I)•d(C) no B

d(IIT)•d(ACC) no B

d(AG)•d(CT) yes B, A

d(AGC)•d(GCT) yes B, A

d(GC) yes B, A, Z

d(GT) yes B, A, Z

d(2AP-T) yes B, A, Z

Effect of Substituents on DNA Conformation

Figure 4-1 Figure 4-9

Figure 1 The fused hexagon motif of A-tract DNA. The four layers are coded by color with the primary layer light blue, the secondary layer magenta, the tertiary layer blue, and the quaternary layer red. The fused hexagon motif is shown in space filling representation, with van der Waal radii of oxygen atoms. (a) Stereoview into the minor groove of the DNA. The DNA is colored by CPK and shown in stick representation. (b) View across the groove, approximately down the normal of the central hexagons. Sites of potassium occupancy are indicated by plus signs. The DNA bases are shaded. Base functional groups that interact with the fused hexagon motif are indicated by circles. (c) The geometry of the sodium form fused hexagon motif. Distances are in red and angles are in white.

Structure of the potassium form of CGCGAATTCGCG: DNA deformation by electrostatic collapse around inorganic cations. Biochemistry. 1998 Dec 1;37(48):16877-87.

http://pubs3.acs.org/acs/journals/doilookup?in_doi=10.1021/bi982063o

Metal cations bind in the minor groove with water

Figure 4-9b

X-ray crystal structure of a 1:1 complex of netropsin: DNA

Netrop_hbond.ppt

The role of minor groove functional groups in DNA hydration. Nucleic Acids Res. 2003 Mar 1;31(5):1536-40.

http://nar.oxfordjournals.org/cgi/content/full/31/5/1536

Loss of O2 carbonyl disrupts spine of hydration

Figure 4-9c

O

O

P

O P

Base

OH O

O

P

O P

Base

OH

C2'-endo, 2E C3'-endo, 3E

B Form of RNA A Form of RNA

Figure. zfactor2.cwg

Z DNA intrastrandphosphate hydration

Z: 6.2 A

Z: 5.6 A

P

O

OH

OH

O

O

P HOH

O

O

P

Conformational and Electrostatic Factors Favoring Various Forms of DNA or RNA

A: 5.7 A

B: 6.7 A

cannotH bond

Figure 4-10

Figure 4-11

A260

molar % dT10 molar % dT10

A260

50 50

1

0.9

0.8

0.7

0.6

File: TAT_mix1.cw2

Evidence for Triplex Helix Formation FromMixing Experiments Monitored by UV

1001000 0 66

Analysis of mixing curves of nucleic acids by UV relies on the hypochromic effect observed upon formation of stacked base apirs

d(T)10 + d(A)10 nd(T)10d(A)10

1

0.9

0.8

0.7

0.6

Figure 4-12

N

N

N

N

O

N

H

H

H

H

H

H

H

H

O

N

N

N

H

H

H

H

H

ONN

N

N

N OO

CH3

H

H

N

N

O

O

CH3

HH

H

H

H

HN

N

N

N

N

Figure: trip_bp1.cw2

Major groove

antiparallel A helix

Watson Crick base pair

parallel

helix,

Hoogsteen

base pair

protonated C required

for base pairing

5 ' - TTTTTTTTTT- 3 '5 ' - AAAAAAAAAA- 3 '3 ' - TTTTTTTTTT- 5 '

HoogsteenWatson-

Crick

5 ' - CCCCCCCCCC- 3 ' ++++++++++5 ' - GGGGGGGGGG- 3 '3 ' - CCCCCCCCCC- 5 '

Hoogsteen

(+ indicates H+)

Watson-

Crick

Polypyrimidine Triplex Motif

Figure 4-13

anti parallelhelix,ReverseHoogsteenbase pair

antiparallel A helixWatson Crick base pair

Major groove

Figure: trip_bp2.cw2

N

H

NN

O

O

CH3

HH

H

H

HHN

N

N

NN

NH

N

N

N

NN

N

N

O

N

H

HH

H

H

HH

H

O

N

NN

H

HN

H

O

NN

N

N

Watson-Crick

ReverseHoogsteen

3'-GGGGGGGGGG-5'5'-GGGGGGGGGG-3'3'-CCCCCCCCCC-5'

Polypurine Triplex Motif

Watson-Crick

ReverseHoogsteen

3'-AAAAAAAAAA-5'5'-AAAAAAAAAA-3'3'-TTTTTTTTTT-5'

Figure 4-14

Intramolecular TriplexNMR structure 1gn7.pdb

Figure triplex.ppt

Figure 4-16

Intramolecular Triplex (1GN7.pdb) highlights

Figure triplex2.pptFigure 4-17

Figure 4-15

5'- AGGAAG GAAGGA 3'3'- TCCTTC CTTCCT 5'

5'- AGGAAG3'- TCCTTC

TCCTTC

3'5'

AGGAAG single strand

triplex

Figure: Hdna1.cw2

GAAGGA

5' 3'

CTTCCT

CTTCCT 3'GAAGGA 5'

single strand

triplex

H DNA, Hoogsteen DNA or Hinged DNAforms in reverse repeat purine DNA under high negative supercoiling

reverse repeatnot inverted repeat

The End Replication Problem

Succesive rounds of replication lead to progressive shortening of the ends of DNA

RNA RNA RNA

missingDNA

replication of this strand results in a shorter DNA

Telomerase solves the End Replication Problem, RNA templated DNA synthesis

elongationelongation

translocation

ribonucleoprotein

Annu. Rev. Pharmacol. Toxicol. 2003. 43:359–79

Figure 4-21b

Schematic structure of a telomere

single strand end protected by DNA displacement loop formation

POTprotectionof telomerebinds TTAG3

The G’s in the telomere sequence can form Quartets via HoogsteenBase Pairing,• Hoogsteen base pairing leads to circluar tetrad.• Center of quartet has large negative electrostatic

potential that can bind cations• all anti glycosyl conformation leads to parallel

stranded quadruplex

Figure 4-22

N

NN

N

O

N

H

H

H

HN

N

N N

O

NHH

H

H

N

NN

N

O

N

H

H

H

H

H

HH

HN

O

N N

NN

Figure: G_tetra1.cw2

+

Four possible orientations of Gn strands

Figure 4-26

parallel antiparallel

mixed (3+1)

antiparallel

Ways of forming intramolecular quadruplexformation with [GxNy]z with 3 types of loops: propeller, lateral, diagonal

Figure 4-27

3'

5'

3'

5'

5'3'

diagonal loop

lateral loop

externalloop

externalorpropellerloop

lateral loop

lateral loop

lateral loop

5'

3'

lateral loop

lateral loop

lateral loop

propeller

basketchair

hybrid

a

aa

a

aa

ss

ss

s s

a

a

a

a

a

a

a

a

a

a

a

a

a

a

a

a

a

a

s

s

ss

s

s

aa

a

a

aa

s

ss

s

s s

Flip centralBase quad

Glycosyl conformation depends on strand orientation. Bases in one base quad can all flip from anti to syn, and syn to anti

Figure 4-28

Front. Chem. 4:38. doi: 10.3389/fchem.2016.00038

J. Phys. Chem. B, Vol. 110, No. 32, 2006 16077

Li+ is strongly hydrated and cannot bindNa+

fits in the plane

K+ fits between the planes

Cramer and Truhlar

propellar

lateral

lateral2

3

syn

anti

A21

T20

T19

G18

G17

G16

A15T14

T13

G12

G11

G10A9

T8

T7

G6

G5

G4

A3

1

1

2

3

G11

G12

T13

T14

A15

G23

G24

T20

A21

T7

G10

A9T8

A3

G4

G5

G6

syn

anti

G22

T19

K+

T8

T7

G4

G5

G10

G11

X

T13A15

G17

G18

T19A21

G12

G16

syn

anti

T20

T14

G6

A9

Y

5'3'

diagonal

lateral lateralT8

T7G10

G11

T13

T14

A15

G17

T19

A21

G12

syn

anti

A9T18

Na+

Na+

Na+

G5

G6G18

G16 G4

laterallateral

diagonal

hybrid-1 hybrid-2

basket form 3

Characterized human telomeric DNA G-quadruplex structures in solution by NMR (See tutorial)

Folding and Unfolding Pathways for the Human Telomeric G-Quadruplex

J. Mol. Biol. (2014) 426, 1629–1650

Nucleic Acids Research, 2016, Vol. 44, No. 22

Single Molecule Fret Studies