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Introduction & applications Part II

Introduction & applications Part II. magnetic tweezers. “MT”. No HW assigned (HW assigned next Monday). Quiz today Bending & twisting rigidity of DNA with Magnetic Traps. Hydrogen.

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Introduction & applications Part II

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  1. Introduction & applications Part II magnetic tweezers “MT” • No HW assigned (HW assigned next Monday). • Quiz today • Bending & twisting rigidity of DNA with Magnetic Traps.

  2. Hydrogen 1. _______________ bonds between neighboring regions of the polypeptide backbone often give rise to regular folding patterns, known as _______________ and _________________. 2. Molecules that assist in the folding of proteins in vivo are called __________________________. 3. Proteins that couple the chemical energy from ATP hydrolysis into mechanical work are called ________________________. 4. A protein generally folds into the shape in which (what function) _________________________ is minimized. 5. At present, the only way to discover the precise folding pattern of any protein is by experiment, using either ______________________ or __________________________________________. α helices β sheets Quiz #4 on (page 119-143) Chpt. 4 of ECB molecular chaperones molecular motors the free energy (G) X-ray crystallography Nuclear magnetic resonance (NMR) spectroscopy

  3. Magnetic Tweezers and DNA Can be conveniently used to stretch and twist DNA. With Super-paramagnetic bead, no permanent dipole. Watch as a function of protein which interacts with DNA (polymerases, topoisomerases), as a function of chromatin: look for bending, twisting. Dipole moment induced, and ma B. t = m x B = 0 U = - m. B F= (m. B) : U ~ -mB2. Δ It is the gradient of the force, which determines the direction, the force is up. (i.e. where B is highest) • DNA tends to be stretched out if move magnet up. • DNA also tends to twist if twist magnets • (since m follows B). • (either mechanically, or electrically move magnets) Forces ranging from a few fN to nearly 100 pN: Huge Range

  4. kB T l F = < x2 > Force measurement- Magnetic Pendulum The DNA-bead system behaves like a small pendulum pulled to the vertical of its anchoring point & subjected to Brownian fluctuations Do not need to characterize the magnetic field nor the bead susceptibility, just use Brownian motion Equipartition theorem: what is it? Each degree of freedom goes as x2 or v2 has ½kBT of energy. Derive the Force vs. side-ways motion. ½ k < x2> = ½ kBT F = k l Note: Uvert. disp = ½ kl2 Udx displacement = ½ k(l2+dx2) Therefore, same k applies to dx . ½ (F/ l) < x2> = ½ kBT T. Strick et al., J. Stat. Phys., 93, 648-672, 1998

  5. Force measurements- raw data kB Tl F = < x2 > Measure < x2 >, l and have F! Measure z, measure dx Find F by formula. Example: Take l = 7.8 mm (4.04 pN-nm)(7800nm)/ 5772 nm = 0.097 pN At higher F, smaller dx; so does dz. Z = l Lambda DNA = 48 kbp = 15 mm At low extension, with length doubling, dx ~ const., F doubles. At big extension (l: 12-14 mm), Dx decrease, F ↑10x. Spring constant gets bigger. Hard to stretch it when almost all stretched out! X T. Strick et al., J. Stat. Phys., 93, 648-672, 1998

  6. The Elasticity of a Polymer (DNA) Chain In the presence of a force, F, the segments tend to align in the direction of the force. Opposing the stretching is the tendency of the chain to maximize its entropy. Extension corresponds to the equilibrium. Point between the external force and the entropic elastic force of the chain. Do for naked DNA; then add proteins and figure out how much forces put on DNA.

  7. Two Models of DNA (simple) Freely Jointed Chain (FJC) & (more complicated) Worm-like Chain (WLC) FJC: Head in one direction for length b, then turn in any direction for length b. [b= Kuhn length = ½ P, where P= Persistence Length] Idealized FJC: Realistic Chain: FJC: Completely straight, unstretchable. No thermal fluctuations away from straight line are allowed The polymer can only disorder at the joints between segments FJC: Can think of DNA as a random walk in 3-D. WLC: Have a correlation length

  8. FJC vs. WLC Bottom line At very low (< 100 fN) and at high forces (> 5 pN), the FJC does a good job. In between it has a problem. There you have to use WJC. FJC WLC

  9. The Freely Jointed Chain Model F b bF Where -F x b cos(q) is the potential energy acquired by a segment aligned along the direction q with an external force F. Integration leads to: And for a polymer made up of N statistical segments its average end-to-end distance is: Langevin Function

  10. Class evaluation 1. What was the most interesting thing you learned in class today? 2. What are you confused about? 3. Related to today’s subject, what would you like to know more about? 4. Any helpful comments. Answer, and turn in at the end of class.

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