Human essential myosin light chain isoforms revealed distinct myosin binding, sarcomeric sorting, and inotropic activity

In this paper, we tested the hypothesis that different binding affinities of the C-terminus of human cardiac alkali (essential) myosin light chain (A1) isoforms to the IQ1 motif of the myosin lever arm provide a molecular basis for distinct sarcomeric sorting and inotropic activity. We employed circular dichroism and surface plasmon resonance spectroscopy to investigate structural properties, secondary structures, and protein–protein interactions of a recombinant head-rod fragments of rat cardiac β-myosin heavy chain aa664–915 with alanine-mutated IQ2 domain (rβ-MYH664–915IQala4) and A1 isoforms [human atrial (hALC1) and human ventricular (hVLC-1) light chains]. Double epitope-tagging competition was used to monitor the intracellular localization of exogenously introduced hALC-1 and hVLC-1 constructs in neonatal rat cardiomyocytes. Contractile functions of A1 isoforms were investigated by monitoring shortening and intracellular-free Ca2+ (Fura-2) of adult rat cardiomyocytes infected with adenoviral (Ad) vectors using hALC-1 or β-galactosidase as expression cassettes. hALC-1 bound more strongly (greater than three-fold lower KD) to rβ-MYH664–915 than did hVLC-1. Sorting specificity of A1 isoforms to sarcomeres of cardiomyocytes rose in the order hVLC-1 to hALC-1. Replacement of endogenous VLC-1 by hALC-1 in adult rat cardiomyocytes increased contractility while the systolic Ca2+ signal remained unchanged. Intense myosin binding of hALC-1 provides a mechanism for preferential sarcomeric sorting and Ca2+-independent positive inotropic activity.