Tools and protocol for quantification of myosin phosphorylation with MRM-MS

Abstract

The phosphorylation of myosin regulatory light chain (LC20) at Thr18 and Ser19 is positively correlated with tension development in smooth muscle tissue, and the molar stoichiometry of LC20 phosphorylation is commonly profiled as a measure of smooth muscle contractility. We provide details for a newly applied multiple reaction monitoring (MRM)-mass spectrometry (MS) method for the quantification of LC20 phosphorylation at Thr18 and Ser19. This MRM-MS method provides a robust alternative to antibody-based detection systems (such as Phos-Tag SDS-PAGE) for the quantification of LC20 phosphorylation. Method name: MRM-MS myosin phosphorylation assay, Keywords: Smooth muscle, Myosin regulatory light chain, LC20, MLC, Artery, Intestine, Targeted proteomics, Quantitation, Stoichiometry, Phosphorylation, Serine 19, Threonine 18