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The structural basis of the cross-bridge cycle. DECORATED ACTIN was the first em reconstruction Moore, Huxley, and DeRosier 1970 J. Mol Biol. Reconstruction by filtered least squares. Projection every 1° (181 projections) Number of observational equations is 161 x 181=29141 - PowerPoint PPT Presentation
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The structural basis of the cross-bridge cycle
DECORATED ACTIN
was the first em reconstruction
Moore, Huxley, and DeRosier
1970 J. Mol Biol.
Reconstruction by filtered least squares
P f = g
The classical solution to the equations
P f= g + e
which minimises the sum of squares oferror seTe is
PT P f = PTg
PT P is th enorma l o r Hessia n matrix
The solution is
f = (PTP)-1 PTg
but PTg is the back projection.
Thus the inverse least squares matrix (PTP)-1
provides the analytical filter for turning theback projection into the required density
Projection every 1° (181 projections)
Number of observational equations is
161 x 181=29141
Order of normal matrix 17645.
Problem should be well determined
Oops!!
Crowther, R. A., DeRosier, D. J. and Klug, A. (1972) The reconstruction of a three-dimensional structure from projections and its application to electron microscopy. Proc. Royal Soc. London 317A: 319-40.
Case 2. Projection every 3°, not enough data to invert the normal matrix. How much data is there?
To determine the rank of a positive (semi) definite matrix we determine its eigenvalue spectrum
Decorated actin at 13Å resolving power calculated by filtered least squares
Electron cryo-microscopy (of decorated actin) shows how strong binding of myosin to actin releases nucleotide Holmes, Angert, Kull, Jahn, Schroeder (2003) Nature 425, 423-427
Four sub-domain refinement
The cross-bridge cycle
As revealed by crystallography
Geeves, M.A. & Holmes, K.C. (2005)
The molecular mechanism of muscle contraction
Adv Protein Chem 71, 161-93
Lymn & Taylor, 1971
The myosin motor domain must move the lever arm from pre- to
post- power stroke.
It must bind to actin strongly or weakly
Both functions are controlled by ATP binding and hydrolysis
Crystallography on the motordomain (particularly Dictyostelium Myosin 2) shows two conformations of myosin:
lever UP and lever DOWN
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Thema 1• Details über das Thema
• Argumente und Beispiele
• Warum ist das Thema für Ihr Publikum von Bedeutung?
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Fischer et al (2005) PNAS102 6873-8
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Fischer et al (2005) PNAS102 6873-8
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Weak and Strong Binding
Coureux, P. D.Wells, A. L.Menetrey, J.Yengo, C. M.Morris, C. A.Sweeney, H. L.Houdusse, A. (2003)
A structural state of the myosin V motor without bound nucleotide
Nature 425 419-23
Rayment_chicken
Upper/lower 50K fitted
Myo 5
?
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