An investigation into the protonation states of the C1 domain of cardiac myosin‐binding protein C

Myosin‐binding protein C (MyBP‐C) is a myofibril‐associated protein found in cardiac and skeletal muscle. The cardiac isoform (cMyBP‐C) is subject to reversible phosphorylation and the surface‐charge state of the protein is of keen interest with regard to understanding the inter‐protein interactions that are implicated in its function. Diffraction data from the C1 domain of cMyBP‐C were extended to 1.30 Å resolution, where the 〈 I /σ( I )〉 of the diffraction data crosses 2.0, using intense synchrotron radiation. The protonation‐state determinations were not above 2σ (the best was 1.81σ) and therefore an extrapolation is given, based on 100% data completeness and the average DPI, that a 3σ determination could be possible if X‐ray data could be measured to 1.02 Å resolution. This might be possible via improved crystallization or multiple sample evaluation, e.g. using robotics or a yet more intense/collimated X‐ray beam or combinations thereof. An alternative would be neutron protein crystallography at 2 Å resolution, where it is estimated that for the unit‐cell volume of the cMyBP‐C C1 domain crystal a crystal volume of 0.10 mm 3 would be needed with fully deuterated protein on LADI III. These efforts would optimally be combined in a joint X‐ray and neutron model refinement.