Ca2 -induced tropomyosin movement in Limulus thin filaments revealed by three-dimensional reconstruction.

1
Ca2-induced tropomyosin movement in Limulus thin
filaments revealed by three-dimensional
reconstruction.

• W. Lehman, R. Craig, P. Vilbert
• Nature, 1994, 386 65-67.

2
Outline

• Background and purpose of paper
• Thin filament background
• Experimental design
• Figures from paper
• Conclusion

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Background / Purpose

• At this point in time, there was some x-ray data,
  but not clear enough to be used for models of the
  thin filament structure.
• Only on state had been resolved by 3D
  reconstruction.
• To show the off and on state of thin
  filaments using Limulus.
• To provide support for steric effects in muscle
  activation.

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Muscle Thin Filament
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Steric Model of Regulated Muscle Contraction
6
Experimental Design

• Used EGTA for off state, Ca2 for on state.
• Negatively stained Limulus thin filaments.
• Constructed density maps of EGTA samples.
• Constructed density maps of Ca2 samples.

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Negatively stained thin filaments

• Limulus thin filaments (in paper)
• Arrows point to bulges ? troponin (in paper)
• A in EGTA
• B in Ca2
• Note this is from a more recent Lehman and
  Craig paper, chicken gizzard thin filaments

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Density Maps
A in EGTA, B in Ca2, C both samples
superimposed
A in EGTA, B in Ca2
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Averaged Layer-Line Data

• Derived from Fourier transforms
• A in EGTA
• B in Ca2
• Note from a more recent Lehman and Craig paper

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Conclusion

• Both the on and off states of troponin in
  native filaments with regulation of troponin can
  be resolved.
• This is direct evidence to support the steric
  model of regulated muscle contraction.