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Abstract:
To obtain high structural resolution for reconstruction of helical objects by single particle
methods a highly accurate alignment along the helical axis is necessary. In case of the
actin-myosin-complex a typical misalignment on the order of the helical symmetry
(±12.7°) limits resolution to 1-2.5nm depending on radius. Applying MSA to
conventionally aligned 2D-projections different projection angles are distinguished. If a
low resolution reconstructed density is available 2D-projections of that density along
different azimuthal angles can be used to define a multi-dimensional factor space. MSA
then defines a 1D-sub-space including its scale for the projection angle. Matching model
data to experimental data for signal strength, resolution, and CTF, these experimental
data can then directly be mapped into the given factor space. The experimental data
shows a close distribution around the sub-space. An azimuthal angle can be assigned by
determining the smallest distance to the sub-space. In a first application using the actinmyosin-
complex it is possible to reduce the error in azimuthal alignment to less than ±3°.
The effect on the final reconstruction and further refinement strategies are discussed.
