Liquid Crystal Phases of DNA and Implications for The Origin of Life

1. Liquid Crystal Phases of DNA and Implications for The Origin of Life Yang Yang, Xianfeng Song Advisor: Sima Setayeshgar Journal Club April 16th, 2008

2. Outline • Part I: Introduction to liquid crystals • Part II: Background on theories of origin of life • Part III: Liquid crystal condensation of 6-to-20-base pair DNA duplexes Liquid crystal condensation of short DNA duplexes provides a means to form long DNA, which is an essential process in origin of life.

3. Part I: Introduction to liquid crystals

4. Introduction to Liquid Crystal • Phases between liquid and solid • Can be divided into two types: • Thermotropic: exhibit phase transition into the LC phase as temperature changed, due to the anisotropy of molecules. • Lyotropic: exhibit phase transition into the LC phase as a function of concentration of the mesogen, formed by amphiphilic molecules in solution. • Mesogen is the fundamental unit of a liquid crystal that induces structural order in the crystals. http://dept.kent.edu/spie/liquidcrystals

5. Birefringence (Double Refraction) • Two different refraction index • no is the refractive indices for o-ray (polarization direction is perpendicular to the optical axis, so called director) • ne is the refractive indices for e-ray (polarization direction is parallel to the optical axis) • A typical behavior due to anisotropy, can be affected by temperature, electrical field, etc • Can twist linear polarization into elliptical polarization if the polarization direction is not parallel or perpendicular to the optical axis. http://plc.cwru.edu/tutorial/enhanced/files/lc/biref/graphics/birefringence.JPG

6. Optical Devices: Crossed Polarizers http://bly.colorado.edu/lcphysics/lcintro/tnlc.html When the polarizers are arranged so that their planes of polarization are perpendicular to each other, the light is blocked. When the second filter (called the analyzer) is parallel to the first, all of the light passed by the first filter is also transmitted by the second. When putting LC in between two polarizers, the polarization state is modified by LC. Now there will be light come through depends on the director’s direction, LC’s thickness, ray’s frequency.

7. Liquid Crystal Phases: Nematic Phase • The mesogens exhibit long-range orientational order • Have no positional order http://dept.kent.edu/spie/liquidcrystals/ From Nature 430, 413-414(22 July 2004) The Schlieren texture, is characteristic of the nematic phase. The dark regions that represent director alignment parallel or perpendicular to the polarizers are called brushes. Schematic of nematic phase

8. Liquid Crystal Phases: Chiral Nematic Phase • The chiral nematic (cholesteric) liquid crystal phase is typically composed of chiral molecules which produces intermolecular forces that favor alignment between molecules at a slight angle to one another. • This leads to the formation of a structure which can be visualized as a stack of very thin 2-D nematic-like layers with the director in each layer twisted with respect to those above and below. http://plc.cwru.edu/tutorial/enhanced/files/lc/phase/phase.htm http://bly.colorado.edu/lcphysics/textures/ A typical texture of chiral nematic liquid crystal with long pitch helix, chracterized by network-like defect lines. The structure of chiral nematic liquid crystals

9. Liquid Crystal Phases: Smectic Phase • Form well-defined layers that can slide over one another • Smectic A phase: the mesogen are oriented along the layer normal • Smectic C phase: the mesogen are tilted away from the layer normal http://plc.cwru.edu/tutorial/enhanced/files/lc/phase/phase.htm Texture of the smectic A phase Picture of the smectic A phase Picture of the smectic C phase

10. Liquid Crystal Phases: Columnar Phases A class of liquid crystal phases in which molecules assemble into cylindrical structures http://www.rsc.org/ej/JM/2001/b008904o/b008904o-f2.gif From Nature 406, 868-871, 2000 From Science 318, 1276 (2007) 100× of texture exhibited by the hexagonal columnar mesophase Columnar phase formed by discotic molecules Columnar phase formed by rod-like molecules

11. Application of liquid crystals • Liquid crystal displays (LCDs): relying on the optical properties of certain liquid crystals in the presence or absence of an electric field. • Thermotropic chiral LCs whoses pitch varies strongly with temperature can be used as crude thermometers, since the color of the material will change as the pitch is changed. LCD display The liquid crystal on the base changes color with temperature change. From http://demo.physics.uiuc.edu/

12. Part II: Introduction to Theories of Origin of Life

13. Origin of Life • Scientific theory • Origin of organic molecules • From organic molecules to protocells • Other theories • Creationism, Spontaneous Generation, Alien Origins (!), …

14. DNA Structure • First X-ray diffraction image of DNA • Taken by Rosalind Franklin in 1952 • Critical evidencein identifying the structure of DNA • Structure model of DNA • Presented by James D. Watson and Francis Crick in 1953 • Double helix with sugar and phosphate parts of the nucleotides forming the two strand • Hydrogen bonds pair nitrogenous bases A and T, C and G • Opposite directions of the two strands of double helix Franklin R, Gosling RG , Nature ,1953 Watson J.D. and Crick F.H.C. Nature, 1953

15. Genetic Information Flow • A gene is a sequence of DNA that encodes for a protein (or groups of proteins) and can influence the phenotype of an organism. The genetic code consists of three-letter 'words' called codons formed from a sequence of three nucleotides (e.g. ACT, CAG, TTT) that encode different amino acids. • In transcription, the codons are copied into messenger RNA (mRNA) by RNA polymerase (RNAP), an enzyme. • In translation, mRNA is decoded to produce a specific polypeptide according to the rules specified by the genetic code.

16. Origin of Organic Molecules • Miller's experiments (Primordial Soup Theory) • The Deep Sea Vent Theory • Wächtershäuser’s Experiment

17. “Miller-Urey” Experiments • Performed by Stanley Miller, and his advisor, Harold Urey in 1953 • Recreates the chemical conditions of the primitive earth in the laboratory • Uses a highly reduced mixture of gases – methane, ammonia and hydrogen – to form basic organic monomers, such as amino acids. • Demonstrates that organic molecules can form under early earth conditions from inorganic precursors Miller S. L., Science ,1953. Miller S. L., and Urey, H. C . Science, 1959 But … How do these relatively simple organic building blocks polymerize and form more complex structures? From NASA

18. Deep Sea Vent Theory • The hot environs of undersea hydrothermal vents were the birthplace for life. • In 1999, Japanese researchers synthesized peptides around an artificial deep-sea vent. Balter, M. Science, 1998 Imai E., et al. ,Science,1999

19. Wächtershäuser's experiment • Early chemistry of life started on mineral surfaces (e.g. iron pyrites) near deep hydrothermal vents • Bubbles on the mineral surfaces acted as the first ‘cell’ • Amino acids and peptides could be formed by mixing simple chemical compounds (carbon monoxide, hydrogen sulfide, nickel sulfide and iron sulfide) Huber, C. and Wächterhäuser, G. , Science, 1998 Wächtershäuser, G. , Science 2000. ajdubre.tripod.com/.../OriginLifeSci-82500.html

20. From Organic Molecules to Protocells • "Genes first" models-the RNA world • "Metabolism first" models-iron-sulfur world

21. RNA World Hypothesis • First put forth by Carl R. Woese in late 1960s • "RNA World” first coined by Gilbert in 1986 • DNA replication need proteins and enzymes while at the origin of life complex proteins are not as yet present • RNA acts as first genetic molecule that can catalyze all the reactions necessary for a precursor to survive and replicate • Relatively short RNA molecules which can duplicate others have been artificially produced in the lab • As complex proteins acting as enzymes are produced, DNA takes the role as the genetic information storage • Woese, C. The Genetic Code, Harper & Row, New York, 1967 • Gilbert, W. Nature, 1986 • Johnston W. K. ,et al. Science, 2001

22. "Metabolism first" Models: Iron-Sulfur World • Early form of metabolism predated genetics • Metabolism here means a cycle of chemical reactions that produce energy in a form that can be harnessed by other processes • Steps for producing proteins: • Produce acetic acid through metallic ion catalysis • Add carbon to the acetic acid molecule to produce three-carbon pyruvic acid (CH3COCO2H) • Add ammonia to form amino acids • Produce peptides and then proteins. • Once proteins are formed, DNA can be replicated and make mRNA, followed by translation to proteins Huber, C. and Wächterhäuser, G. , Science, 1998 Wächtershäuser, G. , Science, 2000.

23. Pending Problem • Once building blocks (small RNAs, small DNAs, small proteins) of a protocell are produced, still need to form larger and more complex ones required by complexity of living organisms. • For example, even one of the simplest organisms, E. coli, has approximately 4,639,221 base pairs in a genome containing 4,403 genes. Human genome contains over 3 billion DNA base pairs. • The formation of long DNA chains by random chemistry predicted by those theories is essentially impossible. The paper gives us an idea how the small molecules tend to self-organize themselves to larger molecule

24. Part III: Liquid Crystal Condensation of sDNA Duplexes

25. Terminology in the Paper • Oligomer: A molecule formed from a small number of monomers. • Self-complementary: Each single strand of the duplex-DNA can form double helix with itself. For example: CCTCAATTGAGG >> <<GGAGTTAACTCC • Non self-complementary: Not self-complementary. For example: CCTCAAAACTCC • sDNA: Short DNA double helix (different than single stranded DNA), with less than 28bps. • lDNA: Long DNA, refers to DNA with more than 28 bps. • DNA ligation: sDNA join together end-to-end to form lDNA.

26. A, B , Z Form of Double strand DNA • B-DNA: Common form in all living creatures. Make a turn every 3.4nm and 10 base pairs. • A-DNA: Similar to B, but with increase in the number of base pairs per rotation. • Z-DNA: Left handed form. Notes form Prof. Cherbas, Dept. of Biology, Indiana University

27. Background on lDNA Liquid Crystal • Duplex lDNA can form liquid crystal phases when hydrated: • Four phases: isotropic phase (I), chiral nematic (N), uniaxial columnar (CU), crystal phase (X) • Ranging from mega base pair (bp) semi-flexible polymers down to approximately 100 bp rigid rod-like segments (B-DNA has bend persistence length ~50nm) • Onsager-Bolhuis-Frenkel (OBF) criterion[*] • Model: Monodisperse repulsive hard rods (length L, diameter D) • Conclusions: If the rods are sufficiently anisotropic in shape, the appearance of nematic phase require: L/D>4.7 (N>28bp). If L/D<4.7, there should be no LC phases at any volume fraction. . [*]Onsager, Ann. N.Y. Acad. Sci. 51, 627 (1949); Bolhuis etc, J. Chem. Phys. 106, 666 (1997)

28. Experiments on sDNA • The solutions which contains a series of self-complementary sDNA duplex-forming “palindromic” oligomers, along with a variety of noncomplementary and partially complementary oligomers • Short complementary B-form DNA oligomers, 6 to 20 base pairs in length, are found to exhibit nematic and columnar liquid crystal phases, even though such duplexes lack the shape anisotropy required for liquid crystal ordering

29. DNA Phase Diagram • The phase diagram includes the phase boundaries measured for sDNA with those obtained from the literature for lDNA, along with the predictions from the Onsager and other models of interacting semi-flexible rod-shaped particle and aggregate solutes. For N < 20, phase transitions from experiments are marked by red open symbols (I-N, triangles; N-CU, circles; CU-C2, squares), and the range of each phase is indicated by colored columns (I, magenta; N, cyan, CU, yellow), at T = 20°C for 20 > N > 8 and T = 10°C for N = 6. • The phase diagram presents evidence that the origin of the LC phases in sDNA is the equilibrium end-to-end physical aggregation of short duplexes into extended duplex units that are long and rigid enough to order. • Estimates of the stacking energy between duplexes are consistent with end-to-end attraction resulting from the hydrophobicity of the faces of their terminal base pairs.

30. LC Ordering from Mixed Solutions of Complementary and Non-complementary Oligomers • In the mixed solutions of complementary and non-complementary DNA oligomers, the complementary ologomers form ridgid duplexes that have a tendency to aggregate end-to-end and form LC phase, whereas the non-complementary oligomers remain as flexible single strands. • Experiment show if there is a large excess of noncomplementary oligomers, the LC phase appears as isolated drops.

31. The observation of nematic and columnar LC phase provides clear evidence for end-to-end stacking of sDNA into rod-shaped aggregates. Discussion & Conclusion • The observation of nematic and columnar LC phase provides clear evidence for end-to-end stacking of sDNA into rod-shaped aggregates. • The end-to-end stacking makes the terminal groups on neighboring oligomers close to each other and the effective concentration much higher than in the surrounding isotropic, thus strongly promoting ligation in the LC phase. • Ligation in the LC phase produces an extended complementary oligomer, further favoring LC transition LC phase acts as the positive feedback for establishing conditions that would strongly promote their own growth into longer complementary chains relative to the non-LC-forming oligomers.

32. Thanks!