Studi sulla muscolatura elicoidale e paramiosinica. III. Osservazioni comparative sulle proteine muscolari di mitilo e oloturia

Ambrosoli Mognoni, Giuseppina   ; Lanzavecchia, Giulio

Atti della Accademia nazionale dei Lincei. Rendiconti della Classe di scienze fisiche, matematiche e naturali, Série 8, Tome 46 (1969) no. 5, pp. 610-618

Résumé

Some proteins obtainable from the primary myofilaments (paramyosinic) of Mytilus muscles (anterior and posterior byssus retractor, and anterior adductor) have been investigated. The proteins have been extracted from myofilaments in a low ionic force medium. No organic solvent has been used. In the crude extracts of the muscle two components have been found: tropomyosin A (93-94%) and another fraction (6-7%) which is prominently myosin. Tropomyosin A was purified by a double crystallization at a low ionic force. The fractions purity was checked by ultracentrifugation (fig. 1) and by gel filtration (fig. 2). In order to prevent the possibility of polymerization at pH7, the physicochemical properties of the tropomyosin A have tested at pH 9; under these conditions, the value of the sedimentation constant was found to be 3.2 (fig. 3), and [$\eta$] was 180 (where c is expressed by g/ml). Tropomyosin A did not show any ATPase activity. At low ionic force it crystallized and crystals showed a period of about 144 Å (Tav. I, fig. 1 c). By using a specific method for the extraction of myosin, we succeded in increasing the concentration of myosin up to 50% (fig. 6); the ATPase activity was always correlated to the myosin concentration. The ATPase activity was higher at pH 9 than at pH 7, and it was more Ca++ than Mg++ dependent. The specific activity of the myosin was about 4000 unities $Q_{P}$; this value corresponds approximately to a number of turnover of 1500, assuming for the myosin a molecular weight of 500.000 (figs. 5 and 7). This specific activity seems to be of the same range of magnitude of that of the Mammals skeletal muscles. These data disagree with Bàràny's papers, according to which the contraction speed of muscles would depend on the ATP activity of myosin. Proteins have been obtained also from longitudinal radial muscles of Holothuria. A protein with similar physico-chemical properties as the tropomyosin A of Mytilus has been extracted. It crystallized at low ionic force, with a period of 144 Å. It was impossible to obtain this protein in the monomeric form, because it had the tendency to polimerize also at pH 9. Nevertheless we could observe some depolimerization (Tav. I, fig. 2) repeating the controls at intervals. It was possible to observe the ATPase activity only in the crude extracts of the muscle. Purified tropomyosin A never showed the ATPase activity. The behaviour of the ATPase activity of the myosin of Holothuria seems to be like that of myosin of Mytilus.

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