Pyrophosphataseについて
スポンサーリンク
概要
- 論文の詳細を見る
In accordance with pH optima four isodynamic pyrophosphatases have been identified. Of these four enzymes, affinities of pyrophosphatase I (pH optimum 4) for pyropho sphate and various phosphate compounds had been reported in 1951. pKm values were 3.21, 4.13, 3.32, 3.00 and 3.31 for pyrophosphate, diphenylpyrophosphate, phenylphosphate, β-glycerophosphate and inorganic phosphate, respectively. In the present investigation, the author purified pyrophosphatase IV (pH optimum 7) of dried yeast and measured the affinities for pyrophosphate and various phosphate compounds by means of direct and indirect methods. As determination of initial velocity calls for the measurement of minute amounts of inorganic phosphate, safranine method was employed. pKm for pyrophosphate was first determined according to Lineweaver and Burk based on the initial velocity of inorganic phosphate production. It was 3.30. pKm for various phosphate compounds, on the other hand, was calculated indirectly, in accordance with the formula of Michaelis and Menten, from the inhibitions of initial hydrolysis of pyrophosphate brought about by the addition of various phosphate compounds. The values of pKm measured by this method were: 2.77, 1.88, 1.72 and 1.98 for diphenylpyrophoshate, phenylphosphate, β-glycerophosphate and diphenylphosphate, respectively. pKm for inorganic phosphate, however, cannot be determined by the addition method as described above. In this case, therefore, the enzyme action was followed until over 50 % ydrolysis of pyrophosphate and the produced inorganic phosphate was estimated by the method of Fiske and Subbrow.Since hydrolytic process would be subjected to competitive inhibition by the decrease in concentration of pyrophosphate (substrate) and the increase of inorganic phosphate, the following formula of Michaelis and Menten was applied to calculate pKi of inorganic phosphate: [numerical formula] When one replaces [numerical formula], respectively, the equation is simplified to [numerical formula] By plotting X against Y, tanθ was graphically obtained; it was 2.06. Consequently the value of Ki, the pKm for inorganic phosphate, was obtained as 3.34 by solving the above formula. It will be seen that the affinities of pyrophospatase I and IV for pyrophosphate and inorganic phosphate are almost similar, but pyrophosphatase IV has smaller affinities for various phosphate compounds than ovrophosohatase I.
- 千葉大学の論文
- 1954-09-28