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Sudipta Maiti Tata Institute of Fundamental Research, Mumbai 30 Years of ASET, TIFR, 18Feb13 Not just Cheaper. Better. Slide 2 Biology is not about cutting frogs anymore: Actually, it never was Robert Hookes microscope Source: Wikipedia Slide 3 There is a revolution on in microscopy Sarkar et al. Frontiers in Membrane Physiology (2012) Label-free Multi-Photon microscopy of serotonin Slide 4 Fluorescence Correlation spectrometer (FCS) Magde et al. (1972) Sengupta et al., Methods (2002) Measuring sub-nm size, inside water Slide 5 Cool Tools You are only as good as your microscope S. Hell (2009) Slide 6 Combining FCS and Multiphoton Kaushalya et al., US Patent no. 7,705,987 (2010) An alignment-free instrument with high sensitivity Slide 7 FCS Workshop 2009 Teaching colleagues from 10 institutes how to build their own Sensitivity > commercially available Cost < 1/8 th Why couldnt they do it before? Slide 8 A culture of building instruments Cutting edge technology is only available in specific labs until it is marketed When are our best labs going to put India on the map of cutting edge scientific instruments? Source: JPK website) Slide 9 Perhaps soon! i2n Technologies, Bangalore Holmarc, Kochi (Technology from TIFR) Slide 10 Abhyankar et al., Proc. SPIE (2012) (Picosecond) (fast) Auto aligned 4 collection Not just cheaper. Better. A dual objective set-up: Half a million photons/sec from a single molecule Slide 11 Why should we do it? A step forward for someone else to build up the knowledge base Recognition from peers all over the world Promote the culture of instrument building among Indian labs and companies Contribution to the economy (?) Slide 12 Thank you Thanks to all my students and collaborators With its extra-ordinary legacy of developing scientific instruments, TIFR MUST TAKE A LEAD Who should do it? Slide 13 Computational Biophysics Group, UIUC Folding intermediates: progress in silico Folding of villin headpiece Experimentalists are far from verifying it Optical: Fast, low resolution, NMR: Slow, high resolution Slide 14 Fragments: concentration can change folding rate X Amyloid aggregation Normal Wolynes and coworkers, PNAS (2013) Separation Unfolding Amyloids: Aggregation and folding are intertwined Self-complimentarity Slide 15 Concentration affects Amyloid- aggregation kinetics Nag et al., J. Biol. Chem. (2011) Oligomers Monomer 150nM 15 nM Why are we interested in A oligomers? Slide 16 Aggregation Number Misfolding Bio-activity Coles et al. Biochemistry (1998) Crescenzi et al. Eur. J. Biochem (2002) Petkova et al. PNAS (2002) 38 DONOR Acceptor Forster Resonance Energy Transfer (FRET) Conformation: Are the oligomers differently folded? Lifetime measures energy transferEnd-to-end distance misfolding Excitation kRkR k NR k Tr Dipole-dipole energy transfer efficiency ~ 1/ R 6 A nanometric ruler for inter- chromophoric distance F rster (1948); Haugland and Stryer (1976) |S 0 > |S 1 > Slide 39 The process preserves the oligomers Before Lyophilization After Lyophilization