Isolation of unstable myosins and the analysis of light chains by capillary electrophoresis

A rapid method for the isolation of unstable fish myosins by Sepharose Q ion-exchange chromatography is described which yields a pure protein essentially free of contamination or breakdown products in less than 6 h. A protocol was developed for determining the molar ratios of myosin light chains (LC) by capillary electrophoresis. The method is quantitative, rapid (<20 min), highly reproducible (<2.1% variation in relative peak migration time), and only uses Femtomole quantities of protein. It was able to separate myosin light chains which could previously only be resolved by 2-D sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Capillary electrophoresis in the presence of SDS gave similar apparent relative molecular masses (M(r)) for most proteins, but an anomalously high M(r) for myosin light chain 3(LC3), as has been reported previously for SDS-PAGE methods.