physics of bio-molecules and cells

CONTENTS Lecturers xi Participants xiii Pr´eface xvii Preface xxi Contents xxv Course 1. Physics of Protein-DNA Interaction by R.F. Bruinsma 1 1 Introduction 3 1.1 Thecentraldogmaandbacterialgeneexpression 3 1.1.1 Two families . . . 3 1.1.2 Prokaryotegeneexpression 5 1.2 Molecularstructure 8 1.2.1 ChemicalstructureofDNA 8 1.2.2 PhysicalstructureofDNA 10 1.2.3 Chemicalstructureofproteins 12 1.2.4 Physicalstructureofproteins 14 2 Thermodynamics and kinetics of repressor-DNA interaction 16 2.1 Thermodynamicsandthelacrepressor 16 2.1.1 Thelawofmassaction 16 2.1.2 Statisticalmechanicsandoperatoroccupancy 19 2.1.3 Entropy,enthalpy,anddirectread-out 20 2.1.4 Thelacrepressorcomplex:Amolecularmachine 23 2.2 Kineticsofrepressor-DNAinteraction 26 2.2.1 Reactionkinetics 26 2.2.2 Debye–Smoluchowskitheory 28 2.2.3 BWHtheory 30 2.2.4 Indirect read-out and induced fit . . . 32 xxvi 3 DNA deformability and protein-DNA interaction 34 3.1 Introduction 34 3.1.1 Eukaryotic gene expression and Chromatin condensation . . 34 3.1.2 A mathematical experiment and White’s theorem . . . . . . 37 3.2 Theworm-likechain 40 3.2.1 CircularDNAandthepersistencelength 42 3.2.2 Nucleosomes and the Marky–Manning transition . . . . . . 42 3.2.3 Protein-DNA interaction under tension 45 3.2.4 Force-ExtensionCurves 47 3.3 TheRSTmodel 50 3.3.1 Structuralsequencesensitivity 50 3.3.2 Thermalfluctuations 52 4 Electrostatics in water and protein-DNA interaction 53 4.1 Macro-ionsandaqueouselectrostatics 54 4.2 Theprimitivemodel 56 4.2.1 Theprimitivemodel:Ion-free 57 4.2.2 Theprimitivemodel:DHregime 57 4.3 Manningcondensation 58 4.3.1 Chargerenormalization 58 4.3.2 Primitivemodel:Oosawatheory 59 4.3.3 Primitivemodel:Freeenergy 61 4.4 Counter-ion release and non-specific protein-DNA interaction . . . 63 4.4.1 Counter-ionrelease 63 4.4.2 Nucleosome formation and the isoelectric instability . . . . 64 Course 2. Mechanics of Motor Proteins by J. Howard 69 1 Introduction 71 2 Cell motility and motor proteins 72 3 Motility assays 73 4 Single-molecules assays 75 5 Atomic structures 77 6 Proteins as machines 78 7 Chemical forces 80 8 Effect of force on chemical equilibria 81 9 Effect of force on the rates of chemical reactions 82 xxvii 10 Absolute rate theories 85 11 Role of thermal fluctuations in motor reactions 87 12 A mechanochemical model for kinesin 89 13 Conclusions and outlook 92 Course 3. Modelling Motor Protein Systems by T. Duke 95 1 Making a move: Principles of energy transduction 98 1.1 MotorproteinsandCarnotengines 98 1.2 SimpleBrownianratchet 99 1.3 Polymerizationratchet 100 1.4 Isothermalratchets 103 1.5 Motorproteinsasisothermalratchets 104 1.6 Designprinciplesforeffectivemotors 105 2 Pulling together: Mechano-chemical model of actomyosin 108 2.1 Swinginglever-armmodel 108 2.2 Mechano-chemicalcoupling 110 2.3 Equivalentisothermalratchet 111 2.4 Manymotorsworkingtogether 112 2.5 Designedtowork 115 2.6 Force-velocityrelation 116 2.7 Dynamical instability and biochemical synchronization . . 118 2.8 Transientresponseofmuscle 119 3 Motors at work: Collective properties of motor proteins 119 3.1 Dynamical instabilities . . 119 3.2 Bidirectionalmovement 120 3.3 Criticalbehaviour 121 3.4 Oscillations . . . . 124 3.5 Dynamic buckling instability . . . . . 125 3.6 Undulation of flagella . . 127 4 Sense and sensitivity: Mechano-sensation in hearing 129 4.1 Systemperformance 129 4.2 Mechano-sensors: Hair bundles . . . . 130 4.3 Activeamplification 131 4.4 Self-tunedcriticality 133 4.5 Motor-driven oscillations . 134 4.6 Channel compliance and relaxation oscillations . . . . . . 136 xxviii 4.7 Channel-driven oscillations 138 4.8 Hearingatthenoiselimit 139 Course 4. Dynamic Force Spectroscopy by E. Evans and P. Williams 145 Part 1: E. Evans and P. Williams 147 1 Dynamic force spectroscopy. I. Single bonds 147 1.1 Introduction 147 1.1.1 Intrinsic dependence of bond strength on time frame forbreakage 148 1.1.2 Biomolecular complexity and role for dynamic force spectroscopy 148 1.1.3 Biochemical and mechanical perspectives of bond strength . 150 1.1.4 Relevant scales for length, force, energy, and time . . . . . . 153 1.2 Brownian kinetics in condensed liquids: Old-time physics . . . . . 154 1.2.1 Two-statetransitionsinaliquid 155 1.2.2 Kineticsoffirst-orderreactionsinsolution 156 1.3 Linkbetweenforce–time–andbondchemistry 158 1.3.1 Dissociation of a simple bond under force . . . 158 1.3.2 Dissociation of a complex bond under force: Stationaryrateapproximation 159 1.3.3 Evolutionofstatesincomplexbonds 163 1.4 Testing bond strength and the method of dynamic force spectroscopy 164 1.4.1 Probemechanicsandbondloadingdynamics 165 1.4.2 Stochastic process of bond failure under rising force . . . . 168 1.4.3 Distributionsofbondlifetimeandruptureforce 169 1.4.4 Crossover from near equilibrium to far from equilibrium unbonding 172 1.4.5 Effect of soft-polymer linkages on dynamic strengths ofbonds 175 1.4.6 Failure of a complex bond and unexpected transitions instrength 177 1.5 Summary 185 Part 2: P. Williams and E. Evans 186 2 Dynamic force spectroscopy. II. Multiple bonds 187 2.1 Hiddenmechanicsindetachmentofmultiplebonds 187 2.2 Impactofcooperativity 188 2.3 Uncorrelatedfailureofbondsloadedinseries 191 2.3.1 Markovsequenceofrandomfailures 191 2.3.2 Multiple-complexbonds 193 xxix 2.3.3 Multiple-idealbonds 194 2.3.4 Equivalentsingle-bondapproximation 195 2.4 Uncorrelatedfailureofbondsloadedinparallel 198 2.4.1 Markovsequenceofrandomfailures 198 2.4.2 Equivalentsingle-bondapproximation 198 2.5 Poissonstatisticsandbondformation 199 2.6 Summary 203 Seminar 1. Polymerization Forces by M. Dogterom 205 Course 5. The Physics of Listeria Propulsion by J. Prost 215 1 Introduction 217 2 A genuine gel 218 2.1 Alittlechemistry 218 2.2 Elasticbehaviour 220 3 Hydrodynamics and mechanics 220 3.1 Motioninthelaboratoryframe 220 3.2 Propulsionandsteadyvelocityregimes 221 3.3 Gel/bacteriumfrictionandsaltatorybehaviour 223 4 Biomimetic approach 225 4.1 A spherical Listeria 225 4.2 Spherical symmetry . . . 226 4.3 Steadystate 227 4.4 Growthwithsphericalsymmetry 229 4.5 Symmetrybreaking 229 4.6 Limitationsoftheapproachandpossibleimprovements 231 5 Conclusion 234 xxx Course 6. Physics of Composite Cell Membrane andActinBasedCytoskeleton by E. Sackmann, A.R. Bausch and L. Vonna 237 1 Architecture of composite cell membranes 239 1.1 The lipid/protein bilayer is a multicomponent smectic phase withmosaiclikearchitecture 239 1.2 The spectrin/actin cytoskeleton as hyperelastic cell stabilizer . . . 242 1.3 Theactincortex:Architectureandfunction 245 2 Physics of the actin based cytoskeleton 249 2.1 Actinisalivingsemiflexiblepolymer 249 2.2 Actinnetworkasviscoelasticbody 253 2.3 Correlation between macroscopic viscoelasticity and molecular motionalprocesses 258 3 Heterogeneous actin gels in cells and biological function 260 3.1 Manipulationofactingels 260 3.2 Control of organization and function of actin cortex by cell signalling . . 265 4 Micromechanics and microrheometry of cells 267 5 Activation of endothelial cells: On the possibility of formation of stress fibers as phase transition of actin-network triggered by cell signalling pathways 271 6 On cells as adaptive viscoplastic bodies 274 7 Controll of cellular protrusions controlled by actin/myosin cortex 278 Course 7. Cell Adhesion as Wetting Transition? by E. Sackmann and R. Bruinsma 285 1 Introduction 287 2 Mimicking cell adhesion 292 3 Microinterferometry: A versatile tool to evaluate adhesion strength and forces 294 4 Soft shell adhesion is controlled by a double well interfacial potential 294 xxxi 5 How is adhesion controlled by membrane elasticity? 297 6 Measurement of adhesion strength by interferometric contour analysis 299 7 Switching on specific forces: Adhesion as localized dewetting process 300 8 Measurement of unbinding forces, receptor-ligand leverage and a new role for stress fibers 300 9 An application: Modification of cellular adhesion strength by cytoskeletal mutations 303 10 Conclusions 303 A Appendix: Generic interfacial forces 304 Course 8. Biological Physics in Silico by R.H. Austin 311 1 Why micro/nanofabrication? 315 Lecture 1a: Hydrodynamic Transport 319 1 Introduction: The need to control flows in 2 1/2 D 319 2 Somewhat simple hydrodynamics in 2 1/2 D 321 3 The N-port injector idea 328 4 Conclusion 333 Lecture 1b: Dielectrophoresis and Microfabrication 335 1 Introduction 335 2 Methods 337 2.1 Fabrication 337 2.2 Viscosity 338 2.3 Electronicsandimaging 338 2.4 DNAsamples 338 3 Results 339 3.1 Basicresultsanddielectrophoreticforceextraction 339 4 Data and analysis 343 xxxii 5 Origin of the low frequency dielectrophoretic force in DNA 347 6 Conclusion 353 Lecture 2a: Hex Arrays 356 1 Introduction 356 2 Experimental approach 360 3 Conclusions 364 Lecture 2b: The DNA Prism 366 1 Introduction 366 2 Design 366 3 Results 367 4 Conclusions 372 Lecture 2c: Bigger is Better in Rachets 374 1 The problems with insulators in rachets 374 2 An experimental test 375 3 Conclusions 381 Lecture 3: Going After Epigenetics 382 1 Introduction 382 2 The nearfield scanner 383 3 The chip 384 4 Experiments with molecules 387 5 Conclusions 391 Lecture 4: Fractionating Cells 392 1 Introduction 392 2 Blood specifics 392 3 Magnetic separation 397 xxxiii 4 Microfabrication 398 5 Magnetic field gradients 399 6 Device interface 401 7 A preliminary blood cell run 406 8 Conclusions 409 Lecture 5: Protein Folding on a Chip 411 1 Introduction 411 2 Technology 412 3 Experiments 415 4 Conclusions 418 Course 9. Some Physical Problems in Bioinformatics by E.D. Siggia 421 1 Introduction 423 2 New technologies 425 3 Sequence comparison 427 4 Clustering 430 5 Gene regulation 432 Course 10. Three Lectures on Biological Networks by M.O. Magnasco 435 1 Enzymatic networks. Proofreading knots: How DNA topoisomerases disentangle DNA 438 1.1 Lengthscalesandenergyscales 439 1.2 DNAtopology 440 1.3 Topoisomerases 441 1.4 Knotsandsupercoils 444 1.5 Topological equilibrium . 446 1.6 Cantopoisomerasesrecognizetopology? 447 1.7 Proposal:Kineticproofreading 448 xxxiv 1.8 Howtodoittwice 449 1.9 Thecareandproofreadingofknots 451 1.10 Suppression of supercoils . . 453 1.11Problemsandoutlook 455 1.12Disquisition 457 2 Gene expression networks. Methods for analysis of DNA chip experiments 457 2.1 Theregulationofgeneexpression 457 2.2 Geneexpressionarrays 460 2.3 Analysisofarraydata 463 2.4 Somesimplifyingassumptions 464 2.5 Probesetanalysis 466 2.6 Discussion 470 3 Neural and gene expression networks: Song-induced gene expression in the canary brain 471 3.1 Thestudyofsongbirds 472 3.2 Canarysong 473 3.3 ZENK 474 3.4 Theblush 476 3.5 Histologicalanalysis 476 3.6 Natural vs. artificial 479 3.7 TheBlushII:gAP 480 3.8 Meditation 481 Course 11. Thinking About the Brain by W. Bialek 485 1 Introduction 487 2 Photon counting 491 3 Optimal performance at more complex tasks 501 4 Toward a general principle? 518 5 Learning and complexity 538 6 A little bit about molecules 552 7 Speculative thoughts about the hard problems 564 Seminars by participants 579 [...]... Bruinsma and Jonathon Howard Physics of Protein-DNA Interaction by Robijn Bruinsma started from the structure of DNA and associated proteins, and lead to discussions of electrostatic interactions between proteins and DNA, and the diffusive search for specific binding sites Joe Howard’s lectures on Mechanics of Motor Proteins discussed mechanical properties of individual proteins and motors, and of complex... Probe Chemistry of Biomolecular Bonds and Structural Transitions explored the rich dynamic behaviors of rupturing individual biomolecular bonds These lectures were followed by Erich Sackmann’s discussion of Micro-rheometry of Actin Networks and Cellular Scaffolds He gave an introduction to membranes and the cytoskeleton and discussed the mechanical properties of cells and the physics of cell adhesion... freedom of motion When i i i i i i i 14 “bruinsma” 2002/8/8 page 14 i Physics of Bio-Molecules and Cells repeated over and over, this reaction produces a flexible string of amino acids – a polypeptide – that starts with an amino group, the so-called “Nterminal” and that ends with a hydroxyl group, the “C-terminal” We saw that the base-pair sequence of the DNA of an organism is a code for the production of. .. relationship between two families of biopolymers: DNA and RNA, constituted of nucleic-acids, and proteins, constituted of amino-acids [1] Proteins are the active agents of the cell As enzymes, they control the rates of biochemical reactions taking place inside the cell They are responsible for the transcription of the genetic code, i.e., the production of copies of short segments of the genetic code that are... bottom-up, so answers to some of the biggest questions about Life are hidden in their smallest parts The 75th Les Houches summer school addressed the physics of biomolecules and cells In biological systems ranging from single biomolecules to entire cells and larger biological systems, it focused on aspects that require concepts and methods from physics for their analysis and understanding The school opened... polypeptide of four amino-acids, starting with an N terminal and ending with a C terminal: Fig 7 Codons and amino-acids (“Ala” stands for “Alanine”, a hydrophobic amino-acid.) Note that only one of the two DNA strands is actually used for the production of proteins, the “coding” strand 1.2.4 Physical structure of proteins The physical structure of protein is determined by two physical mechanisms On the one hand,... for the production of new proteins, and for the duplication of the genetic code, i.e., the production of a full copy of the genetic code during cell division Synthesis of other macromolecules, such as lipids and sugars, is carried out by proteins, the mechanical force of our muscles is generated by specialized proteins adept at “mechano-chemistry”, they detect light, sound, and smell, and maintain the... system of the cell Blueprints for the synthesis of proteins are stored in the form of DNA base-pair sequences, much like strings of zero’s and one’s store information in digital computers A gene is the data string required for the production of one protein (actually, multiple variants of a protein can be produced from the same gene) The beginning and end points of a gene are marked by special “start” and. .. Coli (E.Coli for short) [4] Large numbers of the E.Coli parasitic bacteria live inside your intestines (“colon”) When you drink a glass of milk, part of it will be metabolized i i i i i i i 6 “bruinsma” 2002/8/8 page 6 i Physics of Bio-Molecules and Cells not by you but by your E.Coli bacteria The first step is the breakdown of lactose, sugar molecules consisting of two linked molecular rings Lactose is... breakdown as part of the scheduled maintenance program of the cell) Next, assume that the lactose concentration has dropped The chemical equilibrium now favors the lactose-free conformation of the repressor Lac i i i i i i i 8 “bruinsma” 2002/8/8 page 8 i Physics of Bio-Molecules and Cells repressor binds to the operator sequence and downstream gene transcription is blocked Genetic switches of this type . Micro-rheometry of Actin Networks and Cellular Scaffolds. He gave an introduction to membranes and the cytoskeleton and discussed the mechanical properties of cells and the physics of cell adhesion signalling . . 265 4 Micromechanics and microrheometry of cells 267 5 Activation of endothelial cells: On the possibility of formation of stress fibers as phase transition of actin-network triggered by. 1 ✐ ✐ ✐ ✐ ✐ ✐ ✐ ✐ COURSE 1 PHYSICS OF PROTEIN-DNA INTERACTION R.F. BRUINSMA Department of Physics and Astronomy, University of California, Los Angeles, CA 90024, USA, and Instituut-Lorentz for Theoretical Physics,