抄録
Guanase from human liver was purified, without adding dithiothreitol, into two enzymes, guanase 1 and 2. Studies of these enzymes yielded the following characteristics of guanase.
1. Guanase 1 and 2 showed different substrate (guanine) concentration-activity relationships. Guanase 1 activity displayed typical Michaelis-Menten kinetics, while guanase 2 showed a sigmoid dependence on guanine concentration. Guanase 2 may accelerate catabolism of guanine when the concentration of guanine becomes higher than a fixed value. The Km value of guanase 1 for guanine was 1.78×10-5. Hill plots showed that Hill's n factor for guanase 2 was 1.6 but 1.0 for guanase 1. Therefore, guanase 2 seems to be an allosteric enzyme.
2. 5-Aminoimidazole-4-carboxamide was a competitive inhibitor of guanase, with a Ki of 1.55×10-6. In the presence of 5-aminoimidazole-4-carboxamide, guanase 2 showed typical Michaelis-Menten kinetics. Variation of the concentration of 5-aminoimidazole-4-carboxamide may regulate guanase activity and efficiently accelerate or suppress the catabolism of guanine.
3. Guanase I was inactivated by 10 min treatment of p-chloromercuribenzoic acid at 37°C while approximately 50% of the activity in guanase 2 remained under the same conditions. About 30 to 40% of the guanase 2 activity remained when it was treated with p-chloromercuribenzoic acid up to 30min. Guanase 1 is considered to be inactivated by attack of p-chloromercuribenzoic acid by SH at its active site. In guanase 2, SH in the active site does not seem to be affected by p-chloromercuribenzoic acid because of the difference in its stereo-construction from guanase 1.
4. Al little difference between guanase 1 and 2 was observed in optimum pH and heat stability.
These results indicate that guanase 1 and 2 have different molecular structures.
