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Adenine
Guanine
Cytosine
Thymine
5΄
3΄
DNA Backbone and Bases
BaseO
HO
O
O
P
O
BaseOP OO
O O
O
O
BaseOPO OO
HOCH2Base
O
NN
NN
NH2
N NH
NN
O
NH2
N
N
NH2
O
NH
N
O
O
H3C
(A) (C)
(G) (T)
CC
CC
NN
O
O
H
H
H
CH3
T
3´
5´
CC
CC
NNH
H
O H
H
H
H
NC
NN N
NC
CC
C
HN
CH H
HN
NCC
C C
N
NC
N
O
A
G H
5´
3´
base pairingbetween 2 strands
duplex formation
Watson-Crick pairing
Strand 1 Strand 2
TG
G
T
T
C
A
G C
A
C
A C
G
3´5´
5´ 3´
J.D. Watson, F.H. Crick Nature 1953, 171, 737
W. Fuller, et al
J. Mol. Biol. 1965, 12, 60
(X-ray of DNA fibers at
75% relative humidity)
3 Common Helical Duplexes
A.H.J. Wang, et al
Nature 1979, 282, 680
(CGCGCG in high salt conc.)
A-Form B-Form Z-Form
R. Langridge, et al
J. Mol. Biol. 1960, 2, 19
(X-ray of DNA fibers at
92% relative humidity)
Duplexes in the gas-phase
• V. Gabelica and E. DePauw, Int. J. Mass Spectrom. 2002, 219, 151.
• P.D. Schnier, J.S. Klassen, E.F. Strittmatter and E.R. Williams,J. Am.Chem. Soc. 1998, 120, 9605-9613.
Higher Ea for complimentary duplexes and Ea correlated to –∆Hd in solutionEvidence of Watson-Crick pairing in vacuo
CID yields correlate with number of GC pairs and ∆Hdiss in solutionSuggests structure conserved in gas phase
• M. Rueda, S.G. Kalko, F.J. Luque, M. Orozco J. Am. Chem. Soc. 2003, 125, 8007
Gas-phase MD simulations indicate 12- and 16-mer duplexes retain major
conformational features as the double helix in aqueous solution
Instrumental Details
ESISource
IonFunnel
DriftCell MS Detector
Ion Funnel
DriftCell
Ion Optics
QuadAnalyzer
DetectorTo PumpTo PumpTo Pump
To Pump
Ion Funnel
DriftCell
Ion Optics
QuadAnalyzer
DetectorTo PumpTo PumpTo Pump
To Pump Drift Cell
Ion funnelDetector
Quad
ESIsource
Drift cell
in out
E
~5 torr He
Get shape information from ion mobility
Arrival Time Distribution(ATD)
arrival time
Smaller cross-section
Largercross-section
Molecular Dynamics
Structures Collision Cross-Sections (σ)
ATDs Collision Cross-Sections (σ)
CompareCompare
Experimental Method:
Theoretical Method:
Experiment vs. Theory
H+
Cu+
Na+
Cu+
(2Cu-H)+(3Cu-2H)+
500 600 700 800 900 1000 1100 1200 1300 1400m/z
Duplex
Single-Strand
dCG dCG
dCG+Cu MALDI-TOF Mass Spectrum
Cu+ duplex formation favored over Na+, H+
ATDs for M+(dCG dCG)
1050 1200 1350 1500 1650
arrival time (µs)
Cu+, Ag+
[Zn2+ - H]+ , [Cd2+ - H]+
Li+, Na+, K+
σ expt = 230, 252 Å2
σ expt = 228 Å2[Ca2+- H]+
[Mg2+ - H]+
Cu+
Na+
observed for
[Ni2+- H]+ , [Co2+ - H]+ , [Fe2+- H]+
[Mn2+ - H]+ , [Cr2+- H]+
ATDs for M+(dCG dCG)
1050 1200 1350 1500 1650arrival time (µs)
Ag+
1050 1200 1350 1500 1650arrival time (µs)
Cd2+
Cu+
Zn2+
3d10 4d10
230 Å2
252 Å2257 Å2
226 Å2
227Å2
253 Å2
228 Å2
258 Å2
Theoretical Structure for Na+(dCG dCG)
Na+
PO4 groups
σtheory = 231 Å2
σexpt = 228 Å2
Na+ binds to O on each base
No Watson-Crick pairing
guanine
cytosine
DFT Structures for Cu+(dCG dCG)
Cu+ binds to O on both cytosines
σtheory = 257 Å2
σexpt = 252 Å2
Watson-Crick structureCu+ binds to O on all 4 bases
Cu+
σtheory = 238 Å2
σexpt = 230 Å2
+ ~1 kcal/mol
Summary - metals
d10 metals promote Watson-Crick duplexes, other metals do not
Cu+ > Ag+ >> Zn2+ > Cd2+
dAT Watson-Crick duplexes not observed
[6-mer]3-
[8-mer + Na]4-
[10-mer]5-
[14-mer]7-
658 Å2
σEXPT = 716 Å2
dCG 6, 8, 10, 14dCG 6, 8, 10, 14--mer duplex ATDsmer duplex ATDs
σEXPT = 430 Å2
arrival time
σEXPT = 1000 Å2
536 Å2
σEXPT = 536 Å2
658 Å2
[6-mer]3-
[8-mer + Na]4-
[10-mer]5-
[14-mer]7-
658 Å2
σEXPT = 716 Å2
σEXPT = 430 Å2
arrival time
σEXPT = 1000 Å2
536 Å2
σEXPT = 536 Å2
658 Å2
σGlob = 440 Å2
σGlob = 541 Å2
σGlob = 605 Å2
σGlob = 830 Å2
Theory
dCG 6, 8, 10, 14dCG 6, 8, 10, 14--mer duplex ATDsmer duplex ATDs
σexpt = 430 Å2
[6[6--mer]mer]33-- and [8and [8--mer + Na]mer + Na]44-- Theoretical StructuresTheoretical Structures
σglob = 541 Å2
dCGCGCG(6-mer)
dCGCGCGCG(8-mer)
σglob = 440 Å2
σexpt = 536 Å2
Na+
If the 8-mer duplex is placed in water it remains helical
300K dynamics for 10 ns
Na+
[8[8--mer + Na]mer + Na]44-- 300K Dynamics300K Dynamics
0.5 1 21.5
0.5 1 21.5
0 0.5 1 21.5
Cro
ss-S
ectio
n (Å
2 )C
ross
-Sec
tion
(Å2 )
time (ns)
time (ns)
time (ns)
0
740
720
700
680
660
640
0
Cro
ss-S
ectio
n (Å
2 )
610
710
690
670
650
630
700
680
660
640
600
620
solution helix
solvent-free helix
A-Helix
σexpt = 667 Å2
[8[8--mer + Na]mer + Na]44-- SolventSolvent--Free HelicesFree Helices
B-Helix Z-Helix
σB = 653 Å2 σZ = 654 Å2
7 WC pairs 8 WC pairs
σA = 671 Å2
7 WC pairs
Small amount of the helical form remains in the gas phase on expt time scale
Na+
σexpt = 718 Å2
σA = 734 Å2 σB = 783 Å2
σ = 640 Å2
σZ = 763 Å2
B-Helix Z-HelixA-Helix
globular
[10[10--mer]mer]55-- Theoretical StructuresTheoretical Structures
8 WC pairs 8 WC pairs 10 WC pairs
σexpt = 1002 Å2
σA = 1016 Å2 σB = 1034 Å2
σ = 850 Å2
σZ = 1035Å2
B-Helix Z-HelixA-Helix
globular
[14[14--mer]mer]77-- Theoretical StructuresTheoretical Structures
12 WC pairs 12 WC pairs 14 WC pairs
-90
-60
-30
0
30
60
90
200 220 240 260 280 300 320
λ (nm)
ellip
ticity
(mde
g)
Circular Dichroism Spectrum of the dCG 14-mer in ESI Solution
(50:50 H2O/MeOH, 2%NH4OH)
-90
-60
-30
0
30
60
90
200 220 240 260 280 300 320
λ (nm)
ellip
ticity
(mde
g)
Circular Dichroism Spectrum of the dCG 14-mer in ESI Solution
(50:50 H2O/MeOH, 2%NH4OH)
(ideal)
B-Form
R.R. Sinden, DNA Structure and Function
R.R. Sinden, DNA Structure and Function
-90
-60
-30
0
30
60
90
200 220 240 260 280 300 320
λ (nm)
ellip
ticity
(mde
g)
Circular Dichroism Spectrum of the dCG 14-mer in ESI Solution
(50:50 H2O/MeOH, 2%NH4OH)
(ideal)
B-Form
Z-Form
-90
-60
-30
0
30
60
90
200 220 240 260 280 300 320
λ (nm)
ellip
ticity
(mde
g)
Circular Dichroism Spectrum of the dCG 14-mer in ESI Solution
(50:50 H2O/MeOH, 2%NH4OH)
(ideal)
A-Form
B-Form
Z-Form
J.H. Riazance, et al. Nuc. Acids Res. 1985, 13, 4983
∴ have B form in solution
B-Form A-Form
solvent-free
ESI spray
dehydrate
Ion funnel
dehydrate
solution after spray
~ A-Form
Solution vs. Solvent-Free Structures
k
~ A-Form globular
k is stronglysize dependent
ATDATDdGAGAGAGAGA dGAGAGAGAGA •• dTCTCTCTCTCdTCTCTCTCTC
[10[10--mer]mer]55--
400 500 600 700 800
arrival time (µs)
σexpt = 741 Å2
σexpt = 741 Å2
σA = 750 Å2 σB = 838 Å2
σ = 719 Å2
B-HelixA-Helix
globular
Theoretical StructuresTheoretical StructuresdGAGAGAGAGA dGAGAGAGAGA •• dTCTCTCTCTCdTCTCTCTCTC
[10[10--mer]mer]55--
dTC strand
dGA strand
450arrival time (µs)
550 650 750 850 950
ATD for dATATATATAT duplexATD for dATATATATAT duplex[10[10--mer]mer]55--
IE = 13eV
IE = 44eV
IE = 89eV
758 Å21126 Å2
916 Å2819 Å2
700
750
800
850
900
950
0 100 200 300 400 500 600 700 800 900 1000
dAT 10-mer (A-form) Theoretical Structures
σtheory = 920 Å2
σtheory = 750 Å2σexpt = 819 Å2
σexpt = 916 Å2
σtheory = 820 Å2
σexpt = 756 Å2
time (ps)
cros
s-se
ctio
n (Å
2 )
arrival time (µs)
Assign peaks in ATDAssign peaks in ATD
IE =44eV
dATdAT [10[10--mer]mer]55--
ATD for dCGCGATATATCGCG duplex
arrival time (µs)
IE = 13eV
IE = 44eV
IE = 89eV
450 550 650 750 850 950
1006 Å2
1157 Å2
[14[14--mer]mer]77--
σ = 1006 Å2
σ = 1157 Å2
ATD for dCGCGATATATCGCG duplex
IE = 13eV
1006 Å2
1157 Å2
[14[14--mer]mer]77--
σ = 1006 Å2
σ = 1157 Å2
σtheory = 1000 Å2
(fraying?)
(A or B helix)
dATATCGCGCGATAT ATD
400
arrival time (µs)
500 600 700 800 900
σEXPT = 1123 Å2
[14[14--mer]mer]77--
Summary
• W-C pairing enhanced in dinucleotides cationized by d10 metals
• Duplexes retain helical structures on ms time scale in the gas phase
size dependency – onset of helicity at 8-mer length
base dependency – AT pairs preferentially broken over CG pairs
unidentified conformational family in AT containing
duplexes with σexpt = 1120 – 1160Å2
ManuelDFT calculations
ErinMALDI-TOF
AlessandraESI
JenMD calculations