1

9:05-9:35 4-Feb-2009 Wyss Institute for Biologically Inspired Engineering

Thanks to:

Enabling Technology : Biomaterials Evolution Platform

2

Feb 10

8:30-9:00: Jim Collins - Anticipatory Medical Devices9:05-9:35: George Church - Biomaterial Evolution9:40-10:10: Joanna Aizenberg - Adaptive Architectures10:15-10:45: Don Ingber - Programmable Nanomaterials10:50-11:20: Don Ingber Biomimetic Microsystems

3

Biomaterials Evolution Platform

•DNA Synthesis, assembly & analysis on chips- VDJ-ome (antibody libraries)

• 3-D DNA structures: inorganic interfaces• In vitro systems: multi-enzyme resistance• Microbial systems: multi-virus resistance

- Metabolic engineering fuels & chemicals• Human cell systems: combinatorial regulation

- Organ self-assembly

4

Sequencing tracked Moore’s law 2X/2 yr until 2005-9 10,000x/4yr

2009 goal of 2% for $200 (coding + regulatory)

0.0000001

0.000001

0.00001

0.0001

0.001

0.01

0.1

1

10

1990 1995 2000 2005 2010

$/bp 2005 $50M

2009 $5K

5

Open-architecture hardware, software, wetware

Polonator

$150K - 2 billion beads/run

e.g.1981IBM PC

Rich TerrySenior Engineer

6

DNA Synthesis, assembly & analysis on chips

Nitrobenzyl linkers for selective release of beads or cells

Digital micromirror & inkjet

7

Biomaterials Evolution Platform

•DNA Synthesis, assembly & analysis on chips- VDJ-ome (antibody libraries)

• 3-D DNA structures: inorganic interfaces• In vitro systems: multi-enzyme resistance• Microbial systems: multi-virus resistance

- Metabolic engineering fuels & chemicals• Human cell systems: combinatorial regulation

- Organ self-assembly

8

Antibody (& TCR) VDJ regions

Roth DB et al Mol Cell Biol. 1989 9:3049 N (1-13): 14 22 13 15 10 4 5 4 2 2 3 2 1 Lefranc, The Immunoglobulin FactsBook; Janeway, Immunobiology 2001

VH*DH*NNHH*JJHH*V*JJ

46*23*N N * 6 6 * 67* 55 = > 2M combinations , 750 bp, >1E10 cells

9

Biomaterials Evolution Platform

•DNA Synthesis, assembly & analysis on chips- VDJ-ome (antibody libraries)

• 3-D DNA structures: inorganic interfaces• In vitro systems: multi-enzyme resistance• Microbial systems: multi-virus resistance

- Metabolic engineering fuels & chemicals• Human cell systems: combinatorial regulation

- Organ self-assembly

1010

First practical application of DNA nanostructures

Residual NMR dipolar couplings for membrane proteins >15kDaDouglas, et al. PNAS 2007

11

Selection of DNAs enabling selection of CNT structures

Material properties change at the sub-nm scale (& sensitive to defects) Carbon Nanotubes Tensile strength: 62,000 MPa vs CNT-rope 3600, silk 1000, steel 800Thermal conduction: 6000 W/mK vs Cu 385Electrical current density of 4000 MA/cm2, vs Cu 4

C60 fullerite Bulk Modulus: 491 GPa vs diamond 442, steel 160

12

Biomaterials Evolution Platform

•DNA Synthesis, assembly & analysis on chips- VDJ-ome (antibody libraries)

• 3-D DNA structures: inorganic interfaces• In vitro systems: multi-enzyme resistance• Microbial systems: multi-virus resistance

- Metabolic engineering fuels & chemicals• Human cell systems: combinatorial regulation

- Organ self-assembly

13

Mirror world : resistant to enzymes, parasites, predators

Approach#1: De novo Chemical synthesis (below)

#2: Redesigned peptidyl transferase + D-AA-tRNAs (next slide)

352 AA Synthetic Dpo4 Sulfolobus DNA polymerase IV

4 peptide bonds left to construct

L-aminoacidsD-nucleotides

(current biosphere)

D-aminoacidsL-nucleotides (Mirror-biopolymers)

Duhee Bang

14

113 kbp DNA 151 genes

Pure translation:Forster & Church

MSB ’05 GenomeRes.’06

Shimizu, Ueda ’01

Synthetic LifeHigh speed &

accuracy

Route to mirror proteins

Jewett

15

Biomaterials Evolution Platform

•DNA Synthesis, assembly & analysis on chips- VDJ-ome (antibody libraries)

• 3-D DNA structures: inorganic interfaces• In vitro systems: multi-enzyme resistance• Microbial systems: multi-virus resistance

- Metabolic engineering fuels & chemicals• Human cell systems: combinatorial regulation

- Organ self-assembly

16

Genome Engineering Multiplex Automation (GEMA)

for multi-viral resistance

17

Genome Engineering Multiplex Automation (GEMASS)in vivo homologous allele replacement (lagging SS mimics)

3 hr Cycle time. Application: 314 change for multivirus resistance

18

Improving process yield, health, safety:

What threatens all biological systems?

What do all viruses have in common? or lack?

19

PEG-pAcPhe-hGH (Ambrx) high serum stability314 TAG to TAA changes

IsaacsCharalelChurchSunWang CarrJacobsonKong Sterling

New genetic code: viral-resistance, novel amino acidsno functional GMO DNA exchange

TTT

F

30362 TCT

S

11495 TAT

Y

21999 TGT

C

7048

TTC 22516 TCC 11720 TAC 16601 TGC 8816

TTA

L

18932 TCA 9783 TAASTOP

STOP

2703 TGA STOP 1256

TTG 18602 TCG 12166 TAG 314 TGG W 20683

CTT

L

15002 CCT

P

9559 CAT

H

17613 CGT

R

28382

CTC 15077 CCC 7485 CAC 13227 CGC 29898

CTA 5314 CCA 11471 CAA

Q

20888 CGA 4859

CTG 71553 CCG 31515 CAG 39188 CGG 7399

ATT

I

41309 ACT

T

12198 AAT

N

24159 AGT

S

11970

ATC 34178 ACC 31796 AAC 29385 AGC 21862

ATA 5967 ACA 9670 AAA

K

45687 AGA

R

2896

ATG M 37915 ACG 19624 AAG 14029 AGG 1692

GTT

V

24858 GCT

A

20762 GAT

D

43719 GGT

G

33622

GTC 20753 GCC 34695 GAC 25918 GGC 40285

GTA 14822 GCA 27418 GAA

E

53641 GGA 10893

GTG 35918 GCG 45741 GAG 24254 GGG 15090

1

2

3

4

20

Biomaterials Evolution Platform

•DNA Synthesis, assembly & analysis on chips- VDJ-ome (antibody libraries)

• 3-D DNA structures: inorganic interfaces• In vitro systems: multi-enzyme resistance• Microbial systems: multi-virus resistance

- Metabolic engineering fuels & chemicals• Human cell systems: combinatorial regulation

- Organ self-assembly

21

Accelerated evolution for valuable metabolites & biofuels

22

Biomaterials Evolution Platform

•DNA Synthesis, assembly & analysis on chips- VDJ-ome (antibody libraries)

• 3-D DNA structures: inorganic interfaces• In vitro systems: multi-enzyme resistance• Microbial systems: multi-virus resistance

- Metabolic engineering fuels & chemicals• Human cell systems: combinatorial regulation

- Organ self-assembly

23

Re-programming adult human cells to multiple tissues with combinations of DNAs

Hair or skin sample

24

Viral delivery: β-catenin, Dnmt3l, Dppa2,3,4,5, Ecat1, Eras, Fthl17, Grb21, Ecat8, Fbx15, Gdf3, hTERT, Klf4, Myc, Nanog, Oct4, Rex1, Sall4, Sox2, Sox15, Stat3, Tcl1, Utf1Signals: Activin, BMP4, Estrogen, FGF, HGF, PDGF, Retinoate, Shh, TGF, VEGF.

Combinations of regulatory genes

Cartilage & bone

Respiratory epithelium

Immature Brain

Skin-derived stem cells

25

Biomaterials Evolution Platform

•DNA Synthesis, assembly & analysis on chips- VDJ-ome (antibody libraries)

• 3-D DNA structures: inorganic interfaces• In vitro systems: multi-enzyme resistance• Microbial systems: multi-virus resistance

- Metabolic engineering fuels & chemicals• Human cell systems: combinatorial regulation

- Organ self-assembly

26

.

27

.