[Catalytic properties of purified Ca2+,Mg2+-ATPase from the myometrium sarcolemma]. / Kataliticheskie svoistva ochishchennoi Ca2+,Mg2+-ATPazy sarkolemmy miometriia.

The catalytic properties of myometrium sarcolemmal Ca2+, Mg2(+)-ATPase purified from plasma membrane solubilizate by affinity chromatography on calmodulin-Sepharose were investigated. The enzyme isolated in the presence of azolectin revealed a calmodulin-independent affinity for Ca2+ (Km = 0.17 microM). Purified Ca2+, Mg2(+)-ATPase displayed a strict substrate specificity, was inhibited by low concentrations of o-vanadate and was insensitive to oxytocin and prostaglandins E2 and F2 alpha. The enzyme activity was maximal at 45 degrees C, pH 7.5-8.0, and at Mg-ATP and Ca2+ concentrations of 1.5-2.5 mM and 5-20 microM, respectively.

[Reconstruction of purified Ca2+,Mg2+-ATPase from the myometrium sarcolemma into liposomes and its catalytic properties]. / Rekonstruktsiia ochishchennoi Ca2+,Mg2+-ATP-azy sarkolemmy miometriia liposomy i ee kataliticheskie svoistva.

The Ca2+, Mg2(+)-ATPase of the myometrium sarcolemma purified by the method of affinity chromatography on calmodulin sepharose is reconstituted into azolectin liposomes in the functionally active form by means of cholate dialysis. The ATPase-dependent accumulation of 45Ca is shown on the obtained model system. It makes up 95% of the total accumulation and may decrease to 43% under the effect of 0.8 microM A23187. Ca2+, Mg2(+)-ATPase reconstituted into azolectin liposomes is in the high affinity to Ca2+; Km for Ca2+ is equal to 0.88 +/- 0.22 microM, calmodulin practically does not change it. The highest activity of the reconstituted enzyme is observed at pH 7.0, temperature 50 degrees C, the Mg-ATP concentration 1-2 mM. The Km for substrate is 0.45 +/- 0.02 mM.

[Isolation and purification of Ca2+,Mg2+-ATPase from plasma membranes of the myometrium]. / Vydelenie i ochistka Ca2+,Mg2+-ATP-azy plazmaticheskikh membran miometriia.

The preparation of the purified Ca2+, Mg2(+)-ATPase has been isolated from triton X-100 solubilizate of plasma membranes of the pig myometrium using the method of affinity chromatography on calmodulin-Sepharose 4B. The specific activity of the enzyme shows its 52-fold purification. The enzymic preparation practically has no Mg2(+)-ATPase activity. By the data of DS-Na-electrophoresis in PAAG the Ca2+, Mg2+ ATPase preparation consists of two polypeptides with Mm 130 and 205 kDa. Autoradiography shows their Ca2(+)-dependent phosphorylation. The purified enzyme is highly sensitive to the inhibitory effect of orthovanadate.