Apozymase system

The rate of conversion of coenzyme II to I to some extent followed the degree of fermentation in the apozymase system both processes required hexose diphosphate and were less rapid aerobically than anaerobically. Muscle adenylic acid retarded the conversion. Its rate in the presence of glucose, hexose diphosphate, and phosphate at pH 6.4 (30°C.) calculated from the data of von Euler and Adler (22) is approximately 0.4 m/tM per mg. apozjrmase per hour. 

This conversion has been observed in the apozymase system under conditions similar to those under which the rev<>rse cliangc occurs. It takes place also in an aqueous extract ( maceration juice ) from dried ground bottom yeast, while this is catalyzing the oxidation of hexose diphosphate (1,24). Not any oxidation in which cozymase is involved is adequate for conversion of cozymase to coenzyme II the preparation oxidized alcohol... 

Inactivation of Cozymase in the Pyocyanin System. The usual apozymase system (page 424) will ferment glucose with production of carbon dioxide for some hours. In the presence of fluorides, phosphorylation of glucose still occurred, and this and oxygen uptake were considerably accelerated by pyocyanin (3 X 10 M) but after these additions... 

Reactivation of cozymase -1-0.6 Carbon dioxide evolution in apozymase system 0.075 125... 

The later course of oxygen uptake in the pyocyanin system fell exponentially with time, as did also the rate of cozymase inactivation. This is a reflection of the dependence of the inactivation upon the progress of reactions in w hich cozymase itself plays a necessary part, and whose rate is proportional to the quantity of cozymase available. It can be deduced mathematically and was found experimentally that the total oxygen uptake of such a system is proportional to the quantity of added cozymase. Some 30 moles of oxygen were evolved for each mole of added cozymase. The initial rate of inactivation of cozymase in the reaction mixture of the preceding paragraph was about 3 m/rM per mg. apozymase per hour— which is much more rapid than that of its conversion to coenzyme II (see page 426). 

Inactivation by Apozymase Alone. Inactivation of cozymase was first studied in some detail in the pyocyanin system because it was made evident by the transitory effect of cozymase on oxygen uptake by the system but a modified experimental arrangement showed that the inactivation was brought about by apozymase alone, though with character-... 

More extensive studies were made of th( influence of added substances on the course of cozymase inactivation. This was normally performed at pH 6.2 but was unaffected by variation in pH between 5.6 and 6.8. Nicotinamide, which protects cozymase in other systems (76), did not protect it in apozymase, but the concentration used (5.5 X 10 = M) was relatively low. Increase in phosphate concentration from M/30 to M/6 afforded partial protection, as also did adenosine triphosphate and muscle adenylic acid yeast adenylic acid, adenosine, and adenine were without effect. Hexose diphosphate protected cozymase, and fluoride antagonized the protection as fluoride had no effect on the stability of cozymase in the presence of apozymase alone its effect on cozymase was presumably secondary to the effect on hexose diphosphate breakdown. Of breakdown products from hexose diphosphate, 3-phosphoglyceric acid and 2,3-di-phosphoglyceric acid did not protect cozymase glucose not only did not protect, but also prevented hexose diphosphate from protecting cozymase. 

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