Properties of Microsomal Subfractions Isolated from Developing Rabbit Skeletal Muscle

Abstract

A heterogeneous muscle microsomal fraction isolated from 3–4‐day‐old rabbits was separated into two subfractions by continuous sucrose‐density‐gradient centrifugation. The two subfractions differ from each other morphologically and biochemically.Negative staining reveals that the heavier subfraction (U2) consists of ‘thin‐contour’ vesicles resembling those of sarcoplasmic reticulum membrane from mature muscle. The surface of many of them (about 60%) is covered by 4‐nm particles.The lighter subfraction (U1) contains smooth vesicles with a contour thicker than that of the vesicles of subfraction U2. This difference is probably due to the high lipid content in the vesicles of subfraction U1. The extraction of loosely bound lipids (60% of total lipid content) with anhydrous diethyl ether changes the ultrastructure of these vesicles; their contour becomes thin and the vesicles resemble those in subfraction U2.The activity of Mg2+ ‐dependent ATPase is very high in subfraction U1, but this subfraction lacks calcium uptake and Ca2+‐dependent ATPase activity. Contrary to that, subfraction U2 shows the activity of Ca2+‐dependent ATPase and calcium‐loading capacity in the presence of oxalate.Sodium dodecyl sulphate/polyacrylamide gel electrophoresis shows that the protein pattern of both subfractions is more complicated than that of sarcoplasmic reticulum vesicles from adult muscle. Both subfractions contain proteins of molecular weight between 12000 and 105000. The antibody precipitation technique indicates the presence of Ca2+‐dependent ATPase in subfraction U2 and the presence of calsequestrin in both microsomal subfractions.