Single-Particle Reconstruction Story in a Sample Joachim Frank - - PowerPoint PPT Presentation

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Single-Particle Reconstruction Story in a Sample Joachim Frank - - PowerPoint PPT Presentation

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Single-Particle Reconstruction Story in a Sample Joachim Frank Department of Biochemistry and Molecular Biophysics and Department of Biological Sciences Columbia University 12/08/2017 Nobel Lecture, Stockholm 2 Drawing by Graham Colm,

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Single-Particle Reconstruction Story in a Sample

Department of Biochemistry and Molecular Biophysics and Department of Biological Sciences Columbia University

Joachim Frank

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Nobel Lecture, Stockholm 2 12/08/2017

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Drawing by Graham Colm, courtesy of Wikimedia Commons

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Walter Hoppe (March 21, 1917 – November 3, 1986)

Archives of the Max Planck Society, Berlin

Nobel Lecture, Stockholm 4 12/08/2017

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TO CAPTURE A THREE-DIMENSIONAL OBJECT . . . . . . MULTIPLE VIEWS ARE NEEDED

Nobel Lecture, Stockholm 5 12/08/2017

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Photo credit Brian Clark

Aaron Klug David DeRosier THREE-DIMENSIONAL RECONSTRUCTION: HELICAL SYMMETRY

Nobel Lecture, Stockholm 6 12/08/2017

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THREE-DIMENSIONAL RECONSTRUCTION: VIRUSES WITH ICOSAHEDRAL SYMMETRY Tony Crowther

  • R. A. Crowther, Phil. Trans. Roy. Soc. 1971

Nobel Lecture, Stockholm 7 12/08/2017

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Alpbach, site of workshops on protein X-ray crystallography organized by Walter Hoppe and Max Perutz

Nobel Lecture, Stockholm 8 12/08/2017

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Henderson and Unwin, Nature 1975 THREE-DIMENSIONAL RECONSTRUCTION: TWO-DIMENSIONAL CRYSTAL (PURPLE MEMBRANE PROTEIN)

Nobel Lecture, Stockholm 9 12/08/2017

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Molecule captured in different views Three-dimensional reconstruction produces a 3D image, or density map

Sali et al. Nature 2003 12/08/2017 Nobel Lecture, Stockholm 10

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SINGLE-PARTICLE PROJECTIONS – MOLECULES NEGATIVELY STAINED

Nobel Lecture, Stockholm 11 12/08/2017

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Nobel Lecture, Stockholm 12 12/08/2017

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  • ALIGN & AVERAGE
  • ESTIMATE RESOLUTION
  • SORT/CLASSIFY
  • FIND ANGLES
  • RECONSTRUCT

Nobel Lecture, Stockholm 13 12/08/2017

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Peak

Cross-correlation function Autocorrelation function

  • J. Frank, Ph.D. thesis 1970

Nobel Lecture, Stockholm 14 12/08/2017

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SIMULTANEOUS SHIFT AND ROTATION ALIGNMENT

Nobel Lecture, Stockholm 15 12/08/2017

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2

3

crit

D c dp ≥

PARTICLE SIZE > 3/[CONTRAST2 x RESOLUTION (in Å) x CRITICAL ELECTRON DOSE] CONDITIONS FOR ALIGNMENT OF TWO IMAGES OF A MOLECULE OF SIZE D

Saxton & Frank, Ultramicroscopy 1977

Nobel Lecture, Stockholm 16 12/08/2017

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GLUTAMINE SYNTHETASE

Nobel Lecture, Stockholm 17 12/08/2017

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Zingsheim et al., Proc. Natl. Adad. Sci. USA 1980

ACETYLCHOLINE RECEPTOR

Nobel Lecture, Stockholm 18 12/08/2017

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Frank et al. Science 1981

40S RIBOSOMAL SUBUNIT FROM HELA CELLS

Nobel Lecture, Stockholm 19 12/08/2017

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  • J. Frank, Ph.D. thesis 1970

RESOLUTION = EXTENT OF REPRODUCIBILITY IN FOURIER SPACE BY OPTICAL DIFFRACTION

Nobel Lecture, Stockholm 20 12/08/2017

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Differential Phase Residual

  • J. Frank et al., Science 1981

RESOLUTION = EXTENT OF REPRODUCIBILITY IN FOURIER SPACE USING THE COMPUTER

Nobel Lecture, Stockholm 21 12/08/2017

45°

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Van Heel and Frank, Ultramicroscopy 1981

Limulus polyphemus hemocyanin

Nobel Lecture, Stockholm 22 12/08/2017

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Nobel Lecture, Stockholm 23 12/08/2017

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Nobel Lecture, Stockholm 24 12/08/2017

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Jet Propulsion Laboratory, Pasadena, California

Nobel Lecture, Stockholm 25 12/08/2017

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  • J. Frank, overhead 1979

RANDOM-CONICAL RECONSTRUCTION – PRINCIPLE

Nobel Lecture, Stockholm 26 12/08/2017

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  • J. Frank, American Scientist 1998

RANDOM-CONICAL RECONSTRUCTION – PRINCIPLE (FANCY VERSION)

Nobel Lecture, Stockholm 27 12/08/2017

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Radermacher et al., EMBO J. 1987 RECONSTRUCTION OF 50S RIBOSOMAL SUBUNIT FROM E. COLI RIBOSOME

Nobel Lecture, Stockholm 28 12/08/2017

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SINGLE-PARTICLE PROJECTIONS – MOLECULES EMBEDDED IN ICE

Nobel Lecture, Stockholm 29 12/08/2017

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CONTRAST TRANSFER FUNCTION CORRECTION FROM DEFOCUS SERIES

1

( ) ( ) ( )

N n n n

F W k F

=

=∑ k k

* 2 1

( ) ( ) ( ) ( ) | ( ) | 1

n n n N n n n

SNR k H k W k SNR k H k

=

= +

∑

Penczek et al., Scanning Microscopy 1997

Nobel Lecture, Stockholm 30 12/08/2017

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Frank et al., Nature 1995 Lambert et al., 1994 Radermacher et al., 1994

  • E. coli ribosome Octopus hemocyanin

Calcium Release Channel

Nobel Lecture, Stockholm 31 12/08/2017

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Ban et al., Cell 1998

CRYO-EM RECONSTRUCTION ASSISTING IN PHASING THE X-RAY STRUCTURE

Nobel Lecture, Stockholm 32 12/08/2017

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  • J. Frank, Molecular Machines in Biology, Ch. 2, Cambridge University Press 2011

ANGULAR REFINEMENT (3D PROJECTION MATCHING)

Nobel Lecture, Stockholm 33 12/08/2017

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Valle et al., Cell 2003

RATCHET-LIKE INTERSUBUNIT MOTION Frank & Agrawal Nature 2000

Nobel Lecture, Stockholm 34 12/08/2017

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tRNA AS A MOLECULAR SPRING DURING DECODING

Valle et al., Nat. Struct. Biol. 2003

Nobel Lecture, Stockholm 35 12/08/2017

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HEPATITIS C VIRUS IRES INVADING THE HOST RIBOSOME

Spahn et al., Science 2001

Nobel Lecture, Stockholm 36 12/08/2017

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Villa et al., PNAS 2009

ATOMIC MODELS THROUGH FLEXIBLE FITTING I

Nobel Lecture, Stockholm 37 12/08/2017

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ATOMIC MODELS THROUGH FLEXIBLE FITTING II

Hashem et al., Nature 2013

Nobel Lecture, Stockholm 38 12/08/2017

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Top: classes derived by supervised classification Bottom: classes derived by Maximum Likelihood classification 11,415 particles in common

Scheres et al., Nature Methods 2007

Nobel Lecture, Stockholm 39 12/08/2017

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“STORY IN A SAMPLE”

Sun et al., Nucl. Acid Res. 2015 Adapted from Li et al., Science Adv. 2015

EF-G mutant binding to “PRE” complex Nobel Lecture, Stockholm 40 12/08/2017

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  • T. cruzi ribosome

Liu et al., PNAS 2016 Calcium release channel Des Georges et al., Cell 2017 AMPA receptor Twomey et al., Nature 2017

Nobel Lecture, Stockholm 41 12/08/2017

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Wadsworth Center, Albany, New York State Department of Health Department of Biomedical Sciences, University at Albany Since 2008: Department of Biochemistry and Molecular Biophysics, and Department of Biological Sciences, Columbia University Funding: National Institute of General Medical Science, NIH National Science Foundation Howard Hughes Medical Institute

Nobel Lecture, Stockholm 42 12/08/2017