Quantification of Human Cardiac Ion Channels by Parallel Reaction Monitoring and In-Sample Calibration.

Cardiac ion channels are responsible for action potentials in cardiomyocytes. Dysfunction of ion channels in human hearts by abnormal expression levels causes electrical remodeling in the heart and arrhythmia. Direct quantification of ion channels is scarce because these proteins are large, multipass transmembrane proteins. No targeted proteomic approaches have been carried out to study the copy numbers of these ion channels in human hearts. Following our previous effort to use extended enzymatic digestion on the sodium deoxycholate (SDC) insoluble fractions of the human ventricle to obtain a more comprehensive proteome of sarcomere proteins, we report here the ion channel subunits quantified in copy numbers in the same fraction by a targeted parallel reaction monitoring (PRM) proteomic approach. Using Heavy-Match-Light (HML) in-sample calibration, we successfully quantified three major endogenous ion channel subunits, Nav1.5, hERG, and Cav1.2, and the selected auxiliary subunits, Cavβ1 and Cavα2δ1, in the SDC insoluble fraction of the human ventricle. Our approach showed higher sensitivity than global proteomics for identifying these large membrane proteins. We hope that the developed targeted proteomic method can allow future systematic quantification of these functionally important membrane proteins. We also hope that the absolute quantity obtained from the copy numbers of these proteins allows more comparisons across studies and biological systems, as we demonstrated here for better biological insights.