Connectin, an Elastic Protein of MuscleCharacterization and Function1

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Abstract
An elastic protein, connectin, was prepared from the residues of skeletal and heart muscles from which myosin, actin, and regulatory proteins had been extracted. The residue was further extracted with either 0.1 N NaOH or 1% sodium dodecyl sulfate (SDS), followed by phenol treatment. The yield was approximately 5% of the total myofibrillar proteins in rabbit psoas muscle. Connectin contained about 5% lipids and 1% sugars. The protein had a slightly different amino acid composition depending on whether alkali or SDS treatment was used for extraction. Some connectin was soluble in 1% SDS, but the bulk of the protein did not move onto 10% polyacrylamide gel during electrophoresis, suggesting that the polypeptide chains are covalently crosslinked. After the alkali and phenol treatments, connectin became insensitive to proteolytic enzymes. It was digested by trypsin or elastase, but not by collagenase in myofibrils or in KI-extracted ghost myofibrils. A fiber slack cut from the isolated connectin clump showed a rubber-like elasticity, the Young's modulus being of a similar magnitude to that of unvulcanized rubber. Alkali-treated skinned fibers of frog muscle showed elasticity on passive stretching. However, the tension generation quickly decreased at more alkaline pH than 12. It was constant at pH 11.5. From the tension-sarcomere length relationships of alkali-treated and myosin, actin-extracted skinned fibers and glycerine-treated muscle fibers, it appears that connectin functions as a so-called parallel elastic component of muscle, a component which physiologists have long assumed to exist in order to explain muscle function. Fluorescent antibody against connectin stained whole ghost myofibrils as well as ghost muscle fibers. Electron microscopic observations revealed that the connectin structure is a net consisting of very thin filaments less than 2 nm in diameter. The nets deteriorated upon trypsin treatment, but not upon collagenase treatment. Entanglements of the thin filaments were also observed in thin sections of ghost muscle fibers. In the present study, it is concluded that the connectin structure is responsible for mechanical continuity and tension transmission in striated muscles.