Survey
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
Biopolymers Macromolecules. Macromolecules. Folding of proteins. proteins. Mitotic spindle of a dividing cell November 14th, 2011. Huber Tamá Tamás Actin filament network in the epidermal cell of Tobacco leaf. Biopolymers DNA Polimer: Chains constructed of similar building blocks (monomers, subunits) Number of monomers: tipically 102-104 Titin: 3.435*104 aminoacid C132983H211861N36149O40883S693 Human chromosoma 1: 2.25*108 nucleotide Biopolymer DNA strand released from bacteriophage Subunit 1953 - Rosalind Franklin – DNA double helix X-ray diffraction pattern of DNA Bond Protein Amino acid Covalent (peptide bond) Nucleic acid (RNA, DNA) Nucleotide (CTUGA) Covalent (phosphodiesther) 1953 James D. Watson and Francis Crick – DNA model Polysaccharide (e.g., glycogen) Sugar (e.g., glucose) Covalent (e.g., -glycosidic) T-A (2 hydrogen bonds) Protein polymer (e.g., microtubule) Protein (e.g., tubulin) Secondary C-G (3 hydrogen bonds) DNA primary structure NATURE-1953 Phosphodiester bonds between 2 deoxyriboses 1962 Nobel prize Robert Cecil Olby - The path to the double helix: The Discovery of DNA DNA double helix James D. Watson - The Double Helix Hydrogen bonds between the complementary bases. 1 DNA secondary structure Structure of proteins Primary structure: amino acid sequence Major groove Minor groove Frederick Sanger 1958 Nobel prize Determining the sequence of insuline 1955 A-DNA: short (2.4 nm), wide Hydrogen bonds B-DNA: long (3.4 nm), narrow Electrostatic forces Z-DNA: elongated (alternation of purinepirimidine bases) Van der Waals Secondary structure of proteins Secondary structural elements are stabilized by hydrogen bonds. Peptide bond Tertiary and quaternary structure of proteins Spatial reletionship of secondary structural elements relative to each other. Spatial assembley of subunits β-sheet Hemoglobin α-subunit α-helix Hemoglobin A (2α- and 2β- subunit) Protein folding Driving force of the folding Hydrophobic core Unfolded protein Native state of protein Hydrophil amino acids on the surface 2 Levinthal paradox Anfinsen experiment, RNase A (1961) Wrong conformation Native structure Cyrius Levinthal - 1976 Every peptid unit has ~ 10 conformational states In the case of an 100 aa.-long polipeptide chain 10100 variations id a tio n 1 conformational state: 10-13 s -mercapto ethanol fa st ox Conformation In the reality the folding occurs within slow oxidation Denaturated structure Cyrius Levinthal 1089 s, ~ 1081 years are needed for reaching the native state (longer than the age of the universum) Energy -mercapto ethanol 1 second! Native structure Conclusions: The 3D structure of proteins is determinated by their amino acid sequence. The native structure is thermodynamically the most stabilized state. Energy landscape for protein folding Two-state system General case Misfolded proteins Prion: propageted, misfolded proteins, infectious agents - Transmissible spongiform encephalopaty (Kuru or laughing sickness) Daniel Carleton Gajdusek – 1976 Nobel prize -Creutzfeldt-Jakob disease -bovine spongiform encephalopaty (mad cow disease) Accumulation of β-amyloid – Alzheimer’s disease The depth of the well symbolizes the energetic stabilization of the native state versus the denaturated state. All paths lead to the native state (energetic minimum). Amiloyd plaques in mice brain Models to use to describe the felexibility of polymers Polymer shape resembles random walk (Brownian motion) rN R 2 Nl 2 L LP ri = elementary vector R = ”end-to-end” distance r1 The polymer chain is not rigid, due to its flexibility! “Square-root law”: R Reason of flexibility • • ri LP Diffusion: <x2>=2D <x2> = mean squared displacement D = diffusion constant = diffusion time (duration of observation) = persistence length N = number of elementary vectors Nl = L = contour length • Persistence length informs about the bending stiffnes of the polymer! Members of Fore tribe of New Guinea suffering in Kuru • • 1. Rotation around C-C bonds, 2. Rigid segments connected with flexible (frictionless) joints 3.Torsion of bonds Titin, DNA 1 2 Describing model • 1. Freely Rotating Chain (FRC) • 2. Freely Joint Chains (FJC), • 3. Worm Like Chain (WLC). 3 • Contour length is the streched out length of the polymer! 3 Flexibility of biopolymers Biopolymer mechanics LP=Persistence length L= Contour length Elasticity of the entropic chain Rigid chain LP>>L LP=1-6 mm Entropic elasticity Microtubule Correlation length Semiflexible chain LP~L LP=0.1-20 μm End-to-end distance (R) Actin filament configurational entropy increases (orientation entropy of elementary vectors). Flexible chain LP<<L LP=9-16 nm Force (F) The polymer chain exhibits thermally driven bending motions F = force l = correlation length (persistence length) k B = Boltzmann’s constant T = absolute temperature L = contour length R/L = relative extension Titin Mechanical manipulation of polymers (Optical tweezer) Tying a knot on an single actin filament by optical tweezer! Laser focus DNA molecule Latex bead Highly focused laser beam can trap particles. Moveable micropipette Mechanical manipulation of polymers (Atomic Force Microscopy) Mechanical manipulation of Titin by AFM. Stretching of titin by AFM. F ~ stability of domains Distance between peaks ~ contour length 10μm Deflection of the cantilever can be calibrated as the force used to manipulate the protein. Cantilever is extremely flexible, it deflects under some pN force (10-12) Force extension curve 4 Biological relevance of Titin (Molecular spring) -amiloid filaments on mica surface, scanning mode It consists of over 300 serially linked Ig-domain. Main source of elasticity in the myofibril The End! 5