. The Biological bulletin. Biology; Zoology; Biology; Marine Biology. 328 LEONARD NELSON previously (Nelson, 1955). "Decapitation" was effected by means of a stainless steel, piston-type homogenizer, and the heads and tails separated by repeated gentle centrifugation (1000 X g for ten minutes each) in isotonic KC1. The sedimented heads were discarded. The dilution-precipitation effect was exploited in the further isolation and purification of the tails from the pooled supernatant fractions by the addition of at least five volumes of ice-cold 10~4 M MgCl2. After standing in the refrig
. The Biological bulletin. Biology; Zoology; Biology; Marine Biology. 328 LEONARD NELSON previously (Nelson, 1955). "Decapitation" was effected by means of a stainless steel, piston-type homogenizer, and the heads and tails separated by repeated gentle centrifugation (1000 X g for ten minutes each) in isotonic KC1. The sedimented heads were discarded. The dilution-precipitation effect was exploited in the further isolation and purification of the tails from the pooled supernatant fractions by the addition of at least five volumes of ice-cold 10~4 M MgCl2. After standing in the refrigerator for 6 to 12 hours, the tails had settled out and the clear supernate was decanted and discarded. The flocculent precipitate was further concentrated by centrifugation at 7000 X g for 10 minutes. The sperm tail concentrate was then taken up in two to four volumes of isotonic (M/2) KC1. (All solutions were .40 30 tfi ti +- 3 C 6 o . .10 .0 275 280 285 290 295 300 305 Temperature. (Absolute) 310 315 FIGURE 1. Temperature dependence of Mytilus sperm tail ATP-ase. Reaction mixture: M KG; M Tris buffer, pH ; 10"4 M MgCk: 10'3 M ATP; ml. sperm tail suspension; total volume: ml. Incubation time, 10 minutes. Ordinate, phosphate liberated in 10 minutes; abscissa, temperature, degrees, Absolute. made up in de-ionized water.) Since the tail preparation resisted solubilization in mild alkaline and detergent solutions, no further effort was made to extract them and the experiments were performed on the "intact" tails. The reactants, mixed by lateral agitation in 12-ml. Pyrex conical centrifuge tubes, consisted of or ml. of sperm tail suspension, or 07 ml. Tris buffer (Sigma), pH ( M KC1, 10-4 M MgCL, M Tris), unless otherwise noted. After the reactants had equilibrated in a thermostat at 24° to 25° C, ml. 1Q-3 M ATP (Sigma disodium salt, neutralized with NaOH to bromthymol blue endpoint) was added and the mixture
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