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Phosphorylation-induced structural changes in smooth muscle myosin regulatory light chain
David Kast1, L Michel Espinoza-Fonseca, Christina Yi
1Department of Biochemistry, University of Minnesota, Minneapolis, MN 55455, USA.
Summary
Phosphorylation of the regulatory light chain (RLC) in smooth muscle shifts its structure toward an open state, revealing the initial step in muscle activation. This finding clarifies the RLC phosphorylation mechanism.
Area of Science:
- Biochemistry
- Structural Biology
- Molecular Physiology
Background:
- The phosphorylation domain (PD) of smooth muscle regulatory light chain (RLC) is crucial for muscle contraction but is absent in crystal structures.
- Understanding RLC structural dynamics is key to elucidating smooth muscle activation mechanisms.
Purpose of the Study:
- To elucidate the structural changes in the RLC phosphorylation domain (PD) upon phosphorylation.
- To visualize the atomic-resolution mechanism of smooth muscle activation.
Main Methods:
- Time-resolved fluorescence resonance energy transfer (TR-FRET) experiments on site-directed di-Cys mutants of RLC.
- Molecular dynamics (MD) simulations of RLC bound to myosin subfragment 1 (S1).
Main Results:
- TR-FRET and MD simulations revealed two coexisting structural states of RLC: closed and open.
- Phosphorylation shifts the equilibrium towards the open RLC state by approximately 20%.
- MD simulations provided atomic details of the PD's position and conformation in both states.
Conclusions:
- Phosphorylation of RLC promotes a more helical and extended 'open' state.
- This structural shift is the initial step in the molecular mechanism of smooth muscle activation.
- The combined experimental and simulation approach validates detailed structural insights into RLC regulation.
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