. Cell chemistry; a collection of papers dedicated to Otto Warburg on the occasion of his 70th birthday. Warburg, Otto Heinrich, 1883-; Biochemistry. //M mg/ml Fig. 2. Effect of magnesium con- centration of the rate of acetate- ATP-CoA reaction. Each tube con- tained, in I ml. of reaction mixture: 25 units COA; 10 /j,M ATP; 10 /nM potassium acetate; 200/<Mtris (hydroxymethyl) - aminomethane buffer, pH ; 50 /nM KF; 10 /nM glutathione; and aoo/iMNHjOH, neutralized to pH with KOH; ml (10 units) of yeast fraction 4 and MgClg as indicated. Final pH was The tubes were incu- b
. Cell chemistry; a collection of papers dedicated to Otto Warburg on the occasion of his 70th birthday. Warburg, Otto Heinrich, 1883-; Biochemistry. //M mg/ml Fig. 2. Effect of magnesium con- centration of the rate of acetate- ATP-CoA reaction. Each tube con- tained, in I ml. of reaction mixture: 25 units COA; 10 /j,M ATP; 10 /nM potassium acetate; 200/<Mtris (hydroxymethyl) - aminomethane buffer, pH ; 50 /nM KF; 10 /nM glutathione; and aoo/iMNHjOH, neutralized to pH with KOH; ml (10 units) of yeast fraction 4 and MgClg as indicated. Final pH was The tubes were incu- bated at 37°, and ml. samples were taken at 13, 30, and 60 mi- nutes to determine the rate of the Fig. 3. Effect of pH on the rate of acetate-ATP-CoA reaction. Each vessel contained, in i ml reaction mixture: 25 units CoA; 10 /j,M ATP; 10 /iiM potassium acetate; 50 jiiM KF; 10 f^iM MgClj; 10 /nM glutathione; 200 juM NHjOH (pH adjusted with KOH to that of buffer); 50 fiM each tris (hydroxy- methyl) aminomethane and potas- sium phosphate buffer adjusted to the desired pH; and ml (10 units) of yeast fraction 4. The tubes were incubated at 37°, and ml samples were taken at 20, 40, and 60 minutes to determine the rate of the reaction. neutralized with HCl to pH and the volume adjusted with water to give a CoA solu- tion of the desired concentration. Although solutions store fairly well on freezing, we have routinely prepared fresh solutions each day. CoA concentration. It is a characteristic of the hydroxamicacid system that relatively high concentrations of CoA and hydroxylamine^^ are required for saturation. Under our conditions, as shown in Fig. 4, the maximum activity is approached at about 100 units of CoA. Since the reaction between acetyl mercapto CoA and is non- enzymatic, this relatively high saturation concentration is not surprising. The effect of inorganic phosphate. The addition of 50 to 100 of phosphate per ml generally
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