Impact of methionine oxidation on calmodulin structural dynamics

Area of Science:

• Biophysics
• Structural Biology
• Biochemistry

Background:

• Calmodulin (CaM) regulates target proteins, including the ryanodine receptor (RyR).
• Oxidation of specific methionine residues (M109, M124) in CaM impairs its function.
• Mutations of methionine to glutamine (M to Q) replicate the functional effects of oxidation.

Purpose of the Study:

• To investigate the structural consequences of methionine oxidation in CaM.
• To characterize structural changes induced by M109Q and M124Q mutations using site-directed spin labeling and DEER.
• To understand how these structural alterations affect CaM's calcium-dependent conformational changes.

Main Methods:

• Site-directed spin labeling introduced cysteine residues at T34C (N-lobe) and T117C (C-lobe) of CaM.
• Double electron-electron resonance (DEER), a pulsed EPR technique, measured distances between spin labels.
• Analysis of interspin distance distributions to determine structural states and heterogeneity.

Main Results:

• Ca binding significantly increased the distance between spin labels, consistent with a transition from a closed to an open CaM structure.
• DEER revealed CaM populates both open and closed structural states in the presence and absence of Ca.
• Ca shifts the equilibrium towards the open state by a factor of 13, with each state exhibiting partial disorder.
• M109Q and M124Q mutations reduced the Ca-dependent structural shift, mainly by decreasing the closed-to-open equilibrium constant in the presence of Ca.

Conclusions:

• Methionine oxidation in CaM perturbs its Ca-dependent structural dynamics.
• These structural alterations likely underlie the observed changes in CaM's functional interaction with target proteins like RyR.
• The study highlights the role of methionine oxidation in modulating CaM signaling pathways.