46 INTERMEDIARY METABOLISM AND GROWTH I 



i) "pH 5.2 enzyme" + ATP ^ — r enzyme-AMP ~ PP 



O 



2) Enzyme-AMP ~ PP + RCH(NH2)COOH ^ — , enzyme-AMP ~ C-CH(NH2)R 



+ PP 



^ ?GTP 



3) Microsome-ribonucleoprotein + enzyme-AMP-C-CH(NH2)R > 



microsome-ribonucleoprotein-amino acid-i'tC + AMP + "pH 5.2 enzyme" 



4) ATP + L-methionine "^ — r adenyl-L-methionine + PP 



A protein which catalyzed a methionine dependent pyrophosphate exchange 

 with ATP was also obtained by ammonium sulfate fractionation of the soluble 

 cytoplasmic liver proteins. A similar enzyme system has been reported in yeast 

 (Berg, 1956c). In the presence of methionine, AMP-^''C does not exchange with 

 ATP in the yeast enzyme system. The liver fraction that catalyzes the acyl activa- 

 tion of methionine is similar to the one which also synthesizes "active methionine" 

 (S-adenosylmethionine), see Table 4 reaction 88, p. 36 (Cantoni, 1953). 



K. Glutathione Synthesis 



Enzymes from extracts of acetone dry powders of pigeon liver catalyze the syn- 

 thesis of y-glutamyl cysteine from glutamic acid and cysteine and of glutathione 

 from y-glutamyl cysteine and glycine. ATP is required in both reactions (Snoke 

 and Bloch, 1952; Snoke et al., 1953): 



i) Glutamic + cysteine + ATP ^y-glutamyl cysteine + ADP + H3PO4 



2) Y-Glutamylcysteine + ATP -^ GSH + ADP + H3PO4 



ADP will not substitute for ATP in the above reactions. Both reactions are accel- 

 erated by K^ and Mg"^"". Similar enzymes have been found in wheat germ (Webster 

 and Varner, 1955). The purified enzyme required for the synthesis of glutamyl 

 cysteine does not catalyze an exchange of orthophosphate--^^P with ATP but does 

 catalyze an exchange of AD^^P with ATP. This is consistent with the concept 

 that an enzyme-phosphate compound is an intermediate: 



3) Enzyme + ATP -^ enzyme-P + AD"P 



4) Enzyme-P + glutamic -^ enzyme-glutamic + phosphate 



The enzyme will however catalyze an exchange of orthophosphate in the presence 

 of glutamate. Cysteine reduces the glutamate induced exchange. The enzyme 

 also catalyzes the exchange of cysteine-^^S with y-glutamyl cysteine in the absence 

 of ATP: 



5) Enzyme-glutamate + cysteine -* enzyme + glutamylcysteine 



The mechanism of formation of glutathione from glutamylcysteine and glycine 

 is similar to that for the synthesis of glutamyl cysteine: 



