Single Stranded DNA Topology and DNA Nanotechnology … · Single Stranded DNA Topology and DNA...

Single Stranded DNA Topologyand DNA Nanotechnology

Nadrian C. Seeman

Department of ChemistryNew York University

New York, NY 10003, [email protected]

Lecture NotesAMS Short Course, San Diego

January 05, 2008

DNA is a NanometerScale Object

3.4 Å

~20 ÅB-DNA

DNA is a Double Helix

3.4 Å

~20 ÅB-DNA

The BackboneContains D-SugarsAlternating with

Ionized Phosphates.

.

BACKBONES

DNA

H

(O)H

O

CH H

H

O

H

O

PO O(-)

H

BASE

H

H

O

CH H

H

O

H

O

PO O(-)

H

BASE

H

5'

3'

H

C HH

H

O

H

O

O

PO O(-)

H

BASE

H

H

CH H

H

O

H

O

O

PO O(-)

H

BASE

H

5'

3'

(O)H

(O)H

(O)H

The Backbones areRelated by Twofold

Axes Perpendicular tothe Helix Axis.

.

BACKBONES

DNA

H

(O)H

O

CH H

H

O

H

O

PO O(-)

H

BASE

H

H

O

CH H

H

O

H

O

PO O(-)

H

BASE

H

5'

3'

H

C HH

H

O

H

O

O

PO O(-)

H

BASE

H

H

CH H

H

O

H

O

O

PO O(-)

H

BASE

H

5'

3'

(O)H

(O)H

(O)H

Everything BeginsWith

Hydrogen Bonding

The Watson-CrickBase Pairs

DNA BASE PAIRS

C C

N

C

C

O

NH

O R

H

CH3

T

H

CC

NC

N C

N

N N

H

HC

H

R

A

O

C C

N

N

C

C

R

H

HN

H

H

C

C

NC

C

N

N

C

O

C

H

NR

H

N H

H

G

DNA BASE PAIRS

C C

N

C

C

O

NH

O R

H

CH3

T

H

CC

NC

N C

N

N N

H

HC

H

R

A

O

C C

N

N

C

C

R

H

HN

H

H

C

C

NC

C

N

N

C

O

C

H

NR

H

N H

H

G

How Do We Get TwoBits of Information?

.

NN H

D A

Big

Little

G

C

A

TSize

Size Requires aDouble Helical

Backbone

3.4 Å

Minor Groove

B-DNA

Major Groove

Biological DNA Molecules HaveTwo Components That

Can Be Violated in the Lab:

[1] Watson-Crick Base Pairingand

[2] Regular Double HelicalBackbones

Non-Watson-CrickBase Pairing

Base Triples With

Hoogsteen Base Pairs

H

C C

N

N

C

C

N

NN

H

H

C

H

R

A

CC

N

C

C

O

O

N H

R

H

CH 3

T

T

CH 3

O

O

H

H

N

NC

C

CC

R

Non-Linear DNABackbones in Biology

.

B

Branch

Migrate

Crossover

Isomerize

Resolve

Splic

ePatch

Form

Junction

Ligate

V

v

VI

v i

I I I I I I

I V

A

B

A

B

!

"

!

"

A

B

A

A

B

A A

B

A

B

!

"

!

"

!

"

!

"

!

"

!

"

B

Genetic Recombination

GeneratingNon-Linear DNA

Backbones in DNANanotechnology

Reciprocal Exchange:A Theoretical Biokleptic Tool To Generate

New DNA Motifs

Seeman, N.C. (2001), NanoLett. 1, 22-26.

ReciprocalExchange

Resolve

(Creating a Node by Removing a Zero Node)

(Replacing a Node with a Zero Node)

Reciprocal Exchange in aDouble Helical Context


+

+Strand

Polarity

Identical

Strand

Polarity

Opposite Resolve

Reciprocal

Exchange

Resolve

Reciprocal

Exchange

DS + DS HJ

Biological Reciprocal Exchange:The Holliday Junction

1 4

2 3

1 4

2 3

1 24 3

1 4

2 3

I

I I

A•T

G•C

C•G

C•G

G•C

T•A

A•T

T•A

G•C

T•A

C•G

T•A

A•T

C•G

A•T

G•C

A•T

T•A

G•C

T•A

A•T

C•G

C•G

A•T

G•C

C•G

C•G

G•C

T•A

G•C

A•T

T•A

A•T

T•A

G•C

T•A

C•G

A•T

G•C

C•G

T•A

A•T

C•G

A•T

G•C

T•A

C•G

G•C

A•T

T•A

G•C

T•A

T•A

A•T

C•G

A•T

C•G

A•T

G•C

C•G

C•G

G•C

T•A

G•C

Seeman, N.C. (1982), J. Theor.Biol. 99, 237-247.

Design of Immobile Branched Junctions:Minimize Sequence Symmetry

I IACTCGTGC

TGAGCACG

••••••••

A

T

C

G A

T A

T A

T

C

G

C

G C

G

• • • • • • • •

3322

11 44

C G

C G

CG

A T

A T

AT

CG

C G

•

•

•

•

•

•

•

•

I V

I

I I I

C G

CG

C G

A T

A T

AT

C G

•

•

•

•

•

•

•

C G•

[1] All Short Elements Unique (Tetramers Here).

[2] No Complements to Bend-Spanning Sequences.

[3] No Twofold Sequence Symmetry Flanking the Junction.

[4] Competition with Target 8-mers Only from Trimers.

Highly Branched DNA

C

G•C

G•A

T•

AT•

CG•A

T•

CG •

AT•

C

G• C

G• A

T

•

A

T• C

G•

A

T•

C

G• A

T•

G

C•

C

G•

A

T•

G

C•

G

C•

C

G•

G

C•

A

T•

I I

CG

CG

CG

A

T

AT

AT CGC

G

••

••

•

••

•

AT•

CG•

AT

•

CG

•C

G•AT

•C

G•

C

G•

I

CG •

CG •

AT •

AT •

C G•

A T•

CG •

C G•

C G•

T

T

A

C G

CG

C G

A

A

T

C G

•

•

•

•

•

•

•

I I I I VCG•

CG•

CG•

AT•

C

G•CG•

AT•C

G•

33

11 55

22 44

V

C

G•

C

G•A

T•

A

T•

C

G•

A

T

•

C

G•

A

T•

C

G

•C

G

•C

G•

A

T•

A

T•

A

T•

C

G•

A

T•

JYL5G

22

4433

55

6611

I

CG •

CG •

AT •

AT •

C G•

A T•

CG •

C G•

C G•

T

TA

C G

CG

C G

A

A

T

C G

•

•

•

•

•

•

•

A

T•

C

G

•A

T•

C

G•C

G•A

T•C

G•

C

G•

I I I

C

G• C

G• A

T• A

T•

C

G• A

T

• C

G•

A

T•

C G

C G

CG

A T

A T

AT

CG

C G

•

•

•

•

•

•

•

•

I V

C G•

CG•

C G•

A T•

C G•

C G•

A T•

C G•

•C

G•C

G•T

A•A

TG

C•

•T

A•C

G•T

A•C

GG

C•

•A

T•C

G•C

G•T

AG

C•

•C

G

V

G

C•

C

G•

A

T

•

G

C•

G

C•

C

G•

G

C•

A

T•

I IC

G• C

G

• A

T

•

A

T• C

G•

A

T• C

G• A

T•

C

G•C

G

•C

G•A

T•

A

T•A

T•

C

G•

A

T

•

C

G•

C

G•A

T

•

C

G•

A

T•

A

T

•

C

G•

A

T•

V I

JYL6G

Wang, Y., Mueller, J.E., Kemper, B. & Seeman, N.C. (1991), Biochemistry 30, 5667-5674.

5-Arm and 6-Arm Junctions

Wang, X. & Seeman, N.C. (2007), J. Am. Chem. Soc. 129, 8169-8176.

8-Arm and 12-Arm Junctions

Holding DNA MotifsTogether bySticky Ends

Sticky-Ended Cohesion: Smart Affinity

Qiu, H., Dewan, J.C. & Seeman, N.C. (1997) J. Mol. Biol. 267, 881-898.

Sticky-Ended Cohesion: Structure


The Central Concept of Structural DNA Nanotechnology:Combine Branched DNA with Sticky Ends to

Make Objects, Lattices and Devices

AB'

B

A'

A

B ' B'

B

A

A'

A'

B

OBJECTIVES AND APPLICATIONSFOR OUR LABORATORY

ARCHITECTURAL CONTROL AND SCAFFOLDING

[1] MACROMOLECULAR CRYSTALLIZATION (PERIODIC IN 2D AND 3D).[2] NANOELECTRONICS ORGANIZATION (PERIODIC IN 2D AND 3D).[3] DNA-BASED COMPUTATION (APERIODIC IN 2D OR 3D).[4] CONTROL OF POLYMER AND MATERIALS COMPOSITION & TOPOLOGY.

[1] NANOROBOTICS.[2] NANOFABRICATION.

NANOMECHANICAL DEVICES

SELF-REPLICABLE SYSTEMS

NO

CRYSTALS?PRAY FOR CRYSTALSSET UP CRYSTALS

GUESS NEW CONDITIONS

GUESS CONDITIONS

CHANGE DEITIES

DO CRYSTALLOGRAPHY

YES

CURRENT CRYSTALLIZATION PROTOCOL


A New Suggestion for Producing Macromolecular Crystals

Robinson, B.H. & Seeman, N.C. (1987), Protein Eng. 1, 295-300..

A Method for Organizing Nano-Electronic Components

Robinson, B.H. & Seeman, N.C. (1987), Protein Eng. 1, 295-300.

A Suggestion for a Molecular Memory DeviceOrganized by DNA (Shown in Stereo) Why DNA?

Predictable Intermolecular Interactions:Both Affinity and Structure.

Can Design Shape by Selecting Sequence:Robust Branched Motifs Programmable by Sequence.

Convenient Automated Chemistry:Both Vanilla DNA and Useful Derivatives.

Convenient Modifying Enzymes:Ligases, Exonucleases, Restriction Enzymes, Topoisomerases.

Locally A Stiff Polymer:Persistence Length ~500 Å; Stiff Branched Motifs Have Been Developed.

Robust Molecule:Can Heat Individual Strands without Doing Damage.

Amenable to Molecular Biology and Biotechnology Techniques:Gels, Autoradiography, PCR.

Externally Readable Code when Paired:Different Points in a Lattice Can be Addressed.

High Functional Group Density:Every 3.4 Å Nucleotide Separation.

Prototype for Many Derivatives:The Gene Therapy Enterprise Has Generated Hundreds of Analogs

Potentially Self-Replicable and Selectable:May be Able to Make and Improve Constructs Inexpensively.

DNA Topology Affects DNA Nanoconstructions

Chain Mail Interwoven

What Is the Intellectual Goal ofStructural DNA Nanotechnology?

Controlling the Structure of Matter in 3D to theHighest Extent (Resolution) Possible, so as to

Understand the Connection betweenthe Molecular and Macroscopic Scales.

“What I cannot create, I do not understand.” --Richard P. Feynman

(Inverse not necessarily true.)

STRUCTURAL ANDTOPOLOGICAL

ASSEMBLIES

Polyhedral Catenanes

Cube: Junghuei Chen

Truncated Octahedron: Yuwen Zhang

Chen, J. & Seeman. N.C. (1991), Nature 350, 631-633..

Cube..

Notions Used in Proofof Synthesis=

-

The Cube as the Sum of Belts

Chen, J. & Seeman. N.C. (1991), Nature 350, 631-633..Zhang, Y. & Seeman, N.C. (1994),

J. Am. Chem. Soc. 116, 1661-1669.

TruncatedOctahedron

Notions Used in Proofof Synthesis

1

2 3

4

5

6

I

I I I I I

I V

V

V I

V I I

V I I I

G

A

B

C

D

E

F

Truncated Octahedron Analysis

Zhang, Y. & Seeman, N.C. (1994),

J. Am. Chem. Soc. 116, 1661-1669.

Constructionof

CrystallineArrays

REQUIREMENTS FOR LATTICEDESIGN COMPONENTS

PREDICTABLE INTERACTIONS

PREDICTABLE LOCAL PRODUCT STRUCTURES

STRUCTURAL INTEGRITY

Simple BranchedJunctions are Floppy,like Marshmallows, So

We Return to theMotif Generating

Protocol to Get StiffDX or TX Motifs

Seeman, N.C. (2001) NanoLetters 1, 22-26.

Derivation of DX and TX Molecules

+

+

Resolve

Twice

2 Reciprocal

Exchanges

Resolve

Twice

2 Reciprocal

Exchanges

Resolve

Twice

2 Reciprocal

Exchanges

Resolve

Twice

2 Reciprocal

Exchanges

DS + DS DX TX

Strand

Polarity

Identical

Strand

Polarity

Opposite

Erik Winfree (Caltech)Furong Liu

Lisa Wenzler

2D DX ArraysDA O- O

DA E- O

DA O- E

DA E- E

Topologies of 2D Lattices Made withAntiparallel DX Molecules

Winfree, E. (1996), DNA Based Computing, ed. EJ Lipton, EB Baum. 199-219. Providence: Am. Math. Soc.


Derivation of DX+J Molecules

DX+JDX

+

HP

Resolve

Reciprocal

Exchange

Schematic of a Lattice Containing1 DX Tile and 1 DX+J Tile

16 nm 16 nm

Winfree, E., Liu, F., Wenzler, L.A. & Seeman, N.C. (1998), Nature 394, 539-544.

AFM of a Lattice Containing1 DX Tile and 1 DX+J Tile

Schematic of a Lattice Containing 3 DX Tiles and 1 DX+J Tile

Winfree, E., Liu, F., Wenzler, L.A. & Seeman, N.C. (1998), Nature 394, 539-544.

AFM of a Lattice Containing3 DX Tiles and 1 DX+J Tile

Robust Arrays:DX Triangles

Baoquan Ding

DX Bulged Triangle Motif

Ding, B. & Seeman, N.C. (2004), J. Am. Chem. Soc. 126, 10230-10231.

Two Trigonal Motifs Form aPseudohexagonal Trigonal Array


Zoom

Lattice Views


Zoomed Images


ProgressToward

Three-DimensionalArrays

Chengde MaoJens BirktoftYariv PintoHao YanTong WangRuojie ShaLisa IsraelBaoquan DingBob Sweet

Pam ConstantinouFurong Liu

Phil LukemanJens Kopatsch

Bill ShermanJianping Zheng

Alejandra GaribottiAkinori Kuzuya

Mike Becker

A 3D Trigonal DX Lattice

Chengde MaoJens BirktoftYariv PintoHao YanBob Sweet

Pam ConstantinouPhil Lukeman

Furong LiuBill ShermanMike Becker

X-Ray Diffraction:Predicted Spacings and Symmetry

Resolution: ~10Å

A 3D DNA Tensegrity Triangle[D.Liu, M. Wang, Z. Deng, R. Walulu & C.Mao, J. Am. Chem. Soc. 126, 2324-2325 (2004)]

Designed 142 A Edges

X-Ray Diffraction:Predicted Spacings and

Rhombohedral SymmetryResolution: ~10Å

Large Crystals

These Crystals Singly Extinguish

Under Polarized Light.

Topological Constructions

DNA Knots: John Mueller, Shou Ming Du

RNA Knot & Topoisomerase: Hui Wang

Borromean Rings: Chengde Mao

Caten an e

Knot

Kn ot

Caten an e

STR AN D SW I TCH

STR A N D SW I TCH

Catenanes and Knots are Related

31

A Half-Turn of DNA Corresponds to a Node (Unit Tangle) in a Knot or a Catenane

Seeman, N.C. (1992), Molecular Eng. 2, 297-307.

[-] NODE

RIGHT-HANDED

B-DNA

[-] NODES

[+] NODE

LEFT-HANDED

Z-DNA

[+] NODES

Right-Handed and Left-Handed DNA

DNA Knots

John Mueller

Shou Ming Du

SIN G LE ST R A N D

X X'

Y

Y'

C YC LIC M O LE CU LE

Y'

Y

B -D N A

Z -D NA

X'X'XX

-

-++

C YC LIC M O LEC UL E

B - D N AY

Y'

B - D N A

X'X'XX - -

-


Y'

Y

Z -D NA

Z -D NA

T R EFO IL K N OT (+)

X'X'XX

++

+


X X '

Y

Y '

LIG AT E

M g2 +

0 .1 m M

M g2 +

0 .3 m M

M g2 +

1 0 m M

M g2 +

1 0 m M

)(6

C oN H33++

3 m M3 m M

T R EFO IL K N O T (- )

31-

31+

FIG U R E -8 K N O T

41

T EM PLA T E

1

C I RC LE

0

Mueller, J.E., Du, S.M. & Seeman, N.C. (1991) J. Am. Chem. Soc. 113, 6306-6308.

Du, S.M., Stollar, B.D. & Seeman, N.C. (1995) J. Am. Chem. Soc. 117, 1194-1200.

DNAKnots

-31

+31 4

1

-31

+31 4

1

+ + - + - - - - - -

+

+

+

+

+

+

+

-

-

-

Single-Strand Passages Relate Figure-8Both Trefoil Knots

Du, S.M., Tse-Ding,Y.-C.,. & Seeman, N.C. (1995) Biochem. 34, 673-682.

RNA Knot andTopoisomerase

Hui Wang

-

- -

X X'

Y

Y'S S

SS 16- M ER

DN A

STR A ND

40- M ER

DN A

STR A ND

D N A

LI G ASE

D N A

LI G ASE

D N aseD N ase

RN A S I NGLE STRAN D

RNA K NO TRN A C I RCLE

TO PO I I I

RNAKnot

RNATopoisomerase

Wang, H., Di Gate, R.J. & Seeman, N.C, (1996),

Proc. Nat. Acad. Sci. (USA) 93, 9477-9482.

Borromean Rings

Chengde Mao

Borromean Rings

Mao, C., Sun, W. & Seeman, N.C. (1997), Nature 386, 137-138.

[-] NODE

RIGHT-HANDED

B-DNA

[-] NODES

[+] NODE

LEFT-HANDED

Z-DNA

[+] NODES

General Knot Designs

Seeman, N.C. (1992), Molecular Eng. 2, 297-307.

Junctions, Antijunctions andMesojunctions

Shou Ming DuHui Wang

Half-Turn Motifs Three Half-Turn Motifs

3 421

242

40

44

1

33

Two Half-Turn Motifs

44

403

3

31 4

2

1

242

24 -1 41

Junctions and Antijunctions in a 2D Lattice

A Biological Application:Braiding of

Holliday JunctionsTsu-Ju Fu

Reciprocal Exchange:A Theoretical Biokleptic Tool To Generate

New DNA Motifs


ReciprocalExchange

Resolve

(Creating a Node by Removing a Zero Node)

(Replacing a Node with a Zero Node)

321

Holliday Junction Catenanes

Gel Evidence for Lack of Braiding

4% Denaturing Gel 10% Denaturing GelFROM GENES TO

MACHINES:

DNA NANOMECHANICALDEVICES

PX DNA

Derivation of PX DNA


+

+

DS + DS

Resolve

Everywhere

Reciprocal

Exchange

Everywhere

Resolve

Everywhere

Reciprocal

Exchange

Everywhere

PX

Strand

Polarity

Identical

Strand

Polarity

Opposite

The Real Origin of PX DNA:A Triangle Knot Leading to Switchback DNA

Switchback: The Real Derivation of PX DNA

15'

5'

3'

3'

2 3 4

2

4

3

1

2

4

3

1

2

4

5'5'

3' 3'

A Sequence-DependentDevice

Hao Yan

C D

PX

A B

JX2

A B

D C

Yan, H., Zhang, X., Shen, Z. & Seeman, N.C. (2002), Nature 415, 62-65..

Switchable Versions of PX and JX2

JX2

A B

D C

A B

C D

PX

Machine Cycle of the PX-JX2 Device

A B

C D

A B

C D

A B

D C

JX2

A B

D C

PX

I II

IV III

Switchable Versions of PX and JX2 System to Test the PX-JX2 Device

JX2

JX2

JX2

PXPXPX

AFM Evidence for Operationof the PX-JX2 Device

Yan, H., Zhang, X., Shen, Z. & Seeman, N.C. (2002), Nature 415, 62-65.

RNA Nanotechnologyfrom Luc Jaeger

Hairpin Interactions

Chworos, A., Severcan, I., Koyfman, A. Y., Weinkam,P., Oroudjev,

E. Hansma, H. G., Jaeger, L., Science 306, 2068-2072 (2004).

1D and 2D Structures


E. Hansma, H. G., Jaeger, L., Science 306, 2068-2072 (2004)

Discrete Units


E. Hansma, H. G., Jaeger, L., Science 306, 2068-2072 (2004)

LONG TEMPLATINGSTRANDS

Barcodes fromHao Yan,

Tom LaBean,Lipeng Feng and

John Reif

A 1-D Barcode Assembly

H. Yan, T. H. LaBean, L. Feng & J.H. Reif, Proc. Nat. Acad. Sci. 100, 8103-8108 (2003).

Paranemic Cohesion

Xiaoping Zhang

Paranemic Cohesion with the PX Motif

Left: Ubiquitous Reciprocal Exchange Creates a PX Molecule.Center Right: The Strand Connectivity of a PX Molecule.Far Right: The Blue and Red Dumbbell Molecules are Paranemic.

+

PX Cohesion of DNA Triangles: Theory

A Self-ReplicatableOctahedron from

William Shih,Joel Quispe and

Gerald Joyce

Shih, W.M., Quispe, J.D. & Joyce, G.F., Nature 427,618-621 (2004)

Design for Self-Replicating Octahedron

Shih, W.M., Quispe, J.D. & Joyce, G.F., Nature 427,618-621 (2004)

EM Data for Self-Replicating Octahedron

Flexible Patterningwith Viral DNA

fromPaul Rothemund

Producing Patterns from Long Viral Strands

P.W.K.Rothemund, Nature 440, 297-302(2006).

Various Shapes from Long Viral Strands


Marked Structures and Multimer Patternsfrom Long Viral Strands


Single Stranded DNA Topology and DNA Nanotechnology … · Single Stranded DNA Topology and DNA...

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