Scientia Sinica

No. 1 WANG, TSOU, & WANG: STUDIES ON SUCCINIC DEHYDROGENASE I 81

buffer alone. These results, summarized in Table 6, are in disagreement with the report of Kearney, Singer and Zastrow™,

Table 6, Activity of Succinic Dehydrogenase in Various Buffers.

Buffers Concentration, mM] Other Additions | Concentration, mM Relative Activity Phosphate 50 — — 100 Phosphate 100 — : Saeed 100 Phosphate 150 -— — 105 Phosphate 50 Versene Bar 116 Phosphate 50 Versene 6.9 121 Borate 50 — — 75 Imidazole 50 — — 75 Imidazole 50 Phosphate 5 77 Imidazole 50 Phosphate | 10 80 Imidazole 50 Phosphate | 20 90 Imidazole 50 Phosphate | 50 98 Imidazole 50 Phosphate 90 103 Imidazole 50 Versene . Bua) 102 Imidazole 50 Versene 6.9 105 Imidazole 50 Versene Oe: 104

ee ea eEeenaSnnna nnn

The specific activity of the enzyme preparation used in these experiments was 76. During enzymic asszy in borate or imidazole buffer, the enzyme was-first dialyzed against imidazole buffer. The dialyzed enzyme had a phosphate concentration by direct determination of 1.55 mM. As 0.1 mi enzyme was used per 4 ml reaction mixture, the final phosphate concentration was only 38 uM. .

Effect of succinate concentration. Fig. 4A shows the effect of succinate concentration on enzyme activity. It can be seen that the enzyme is nearly saturated with its substrate at a succinate concentration of 2 10° M. Line weaver and Burk”! plots give Michaelis constants of 1.9 and 0.58 mM at 0° and 38°C respectively (Fig. 4B). The latter value is in essential agreement with the values of 0.25-0.51 mM, reported by Thorn! using an assay method very similar to ours.

Effect of different acceptors. With ferricyanide as acceptor, the reaction rate increases with increasing ferricyanide concentration until a concentration of 5 mM is reached, beyond which further increase in ferricyanide becomes inhibitory. Within this concentration range, a plot of the reciprocals of the reaction rates against those of ferricyanide concentrations gives a straight line

(Fig. 5).