Forces organizing soft matter: large bio-molecules, colloids, polymers

Transcription

1 Forces organizing soft matter: large bio-molecules, colloids, polymers * Theory and measurement of intermolecular forces * Single molecule transport driven by molecular interaction Visiting scientist: Rudi Podgornik (course Physics of DNA!) Grad Students: Selcuk Yasar, Jaime Hopkins, Alphan Aksoyoglu, Brian Hildebrandt Undergrads: Adam Cohen, Andrew Clark

2 Theory of intermolecular forces (electrostatics, electrodynamics, solvation, steric) example: single-walled carbon nanotubes Connection between dielectric spectra and van der Waals torques/forces. Dept of Energy review: French, Parsegian, Podgornik et al. Rev Mod Phys 82: Long Range Interactions Now under under study: DNA, Lipids (cell membranes), Proteins Rudi Podgornik, course, Physics of DNA

3 Measurement of forces and energies: example DNA-DNA repulsion Equation of State: Osmotic pressure vs. DNA density, function of salt, temperature DNA inside virus A. Evilevitch

4 Single-molecule transport: example, ionic channels E(mV) Detection _ + I(pA) pa pa 0 25 µm 10 ms 10 ms 1 cm Current through channel 70 µm Sergey Bezrukov, Philip Gurnev 100 µm 2 nm

5 Unified equation of state for flexible polymers: Osmotic pressures of many sizes and concentrations fit with ONE parameter. Now measuring pressures in mixed systems, seeing polymers push other polymers through channels.

6 Ross Lab: Biophysics of Microtubules and Motors Cytoskeleton: Rich Biological System of Interest 1.Microtubules shape the cell and remodeling of the network is important biological and intriguing physical problem a.biology: cell division, cell differentiation, development b.physics: Non-equilibrium processes and selforganization. 2.Mechanics and dynamics are physical parameters important for cellular activities. 3.Amenable to high-resolution single molecule microscopy methods Website:

7 Ross Lab - Biophysics Current Projects: 1.Biophysics/biochemistry of microtubule severing enzymes 2.Measuring flexural rigidity of microtubules with associated proteins bound 3.Self-organization and nanoscale traffic of microtubule motors 4.Building and characterizing the FPALM/STORM microscope Diaz-Valencia, et al. Biophysical Journal May, My lab is fun and full. Always someone to talk to. Two postdocs 2. You get to learn biology. More job options than physics alone. 3. We are writing papers! Website:

8 Ross Lab - Biophysics Methodologies: 1.Biochemistry and Molecular Biology: Protein Design and Engineering, Purification, and Characterization 2.Biophysical methods: In vitro Reconstitution Assays, Bottom-up Engineering of Cellular Systems, 3.Microscopy: Epi-fluorescence, TIRF, electron microscopy, AFM 4.Analysis: Image analysis, MATLAB analysis, measure velocity, intensity, Fourier modes Website:

9 Dynamics and mechanics of a range of biomaterials Kilfoil Lab Mechanotransduction in cell membranes Colloids as a model biomaterial Dynamics of cell division A bottom-up approach to cell mechanics 17µ m l p κ k T B l p mm Scale bar 10 µm

10 Force generation and transduction in these cell materials Interphase Prophase Prometaphase Metaphase Anaphase Telophase L(t) (µm) movie: Saccharomyces cerevisiae expressing fluorescent labels: poles of mitotic spindle (spindle pole spb separation 2nd centromere detection bodies, or SPB) chromosomes at attachment site (centromere) t (min)

11 Towards more complex in vitro cell mechanics models!"# Poisson s ratio "!!!"#!$ $! $ $! ) r%,!-. $!!+ F-actin 1mg/ml D rr &r,!(%,!- ). Composite MT + F-actin network $!!* $!!#!!"# $ $"#!%&'( Poisson s Ratio of Material 1 mg/ml actin 2.4 mg/ml microtubules Scale bar 10 µm 1.0 µm beads 1.9 µm beads Close-up: axes in µm 162 µm Random walk of single bead

12 Kilfoil Lab: Dynamics and mechanics of a range of biomaterials Methodologies Microscopy: 3-Dimensional confocal microscopy, multi-point scanner Analysis: Develop new image analysis methods, data analysis in vivo expression of fluorescent proteins and genetic manipulation of cells: Microfluidics Biophysical methods: In vitro reconstitution of protein networks

13 Single Biomolecules in Confining and Crowded Environments Lori S. Goldner Graduate students: Peker Milas Richard Buckman Ben Gamari Sheema Rahmanseresht Nina Zefroosh (summer 2011) Overall Objectives: Understand the physics of biological molecules and molecular complexes in confining or crowded environments that mimic the environment of the cell. Specific Projects or Interests: Short DNA flexibility Short RNA/DNA interactions with proteins. Molecular assembly. Protein assembly. Cellulose synthesis. Approach: Develop and apply novel optical techniques for single molecule detection, confinement, and manipulation. R.V. Miller, Scientific American 1998

14 Biomolecules in Nanodroplets We use nanodroplets both to facilitate measurement of biomolecular interactions, and to enhance relevance to real biological systems. Appl. Phys. Lett. 89, (2006). 2

15 Antisense Interactions in RNA One strand of RNA binds to a complementary region of another strand of RNA in an interaction that can be regulatory or enzymatic in function. Often a kissing or loop-loop interactions such as that shown on the left is involved. Kissing interactions are often the first step in a structural change such as that shown on the right. 3

16 Cellulose synthesis (with Tobias Baskin, Biology) Cellulose synthesizing complexes (CSC) in the membrane of a plant cell. Image taken from Delmer, D. P Annual Review of Plant Physiology and Plant Molecular Biology (1999). Brownian ratchet model of cellulose synthesis. From Diotallevi, F. and Mulder, B. Biophys. J. (2007). Image of fluorescently-labeled CESA-3 in living plant cells, portion of three cells shown. Image taken by Nina Zefroosh in the Goldner lab. 4

17 Bacterial locomotion: swarming Swarming is high-density motion of bacteria on a surface The exterior portion of a swarm colony self-organizes into a single-cell-thick layer This is a naturally occurring biological analog of the Menon group s system of vibrated rods. The swarm is naturally modeled as a 2D self-propelled gas We have measured local short-range correlation functions, on the order of the cell length. We have observed longer-range swirling, but not yet characterized it. We ultimately want to understand bulk flow rates, local jamming and multiple layer formation. Swarming E. coli. A typical cell is 6 µm long. Correlation between cells velocities.

18 Bacterial locomotion: flagellar polymorphism Bacteria swim by rotating helical flagella Flagella occasionally change shape during swimming. The transformation between different polymorphic forms is triggered by changes in the torque being applied by the flagellar rotary motor. The shapes themselves are understood but not the energy associated with shape change. Each shape corresponds to a different fraction of the flagellin proteins switching to a higher-free-energy state with more inherent twist. Swimming E. coli with fluorescently labeled flagella. Repeatedly pulling on a single flagellum in an optical trap gives a force-displacement curve. Smooth portions of the curve are elastic stretching of the helix A vertical jumps occurs when a section of helix transforms from one polymorphic form to another. Using the statistics of the transition between forms, we will measure the relative thermodynamic stability of different forms. Flagellum stretched in an optical trap.

19 Thanks