IV. BIOCHEMICAL SYSTEMS 31 



triphosphate was indispensable for the methylation of GA by methionine 

 if homogenized Uver preparations were employed. i-^' ^^° In this case, oxygen 

 was also essential, and the reaction was affected by inhibitors of oxidative 

 mechanisms. Cohen^^^- "- showed that homogenized liver preparations of 

 many species, including cattle, sheep, swine, chickens, hamsters, and 

 rabbits, formed little or no creatine in the presence of GA, methionine, 

 ATP, and oxygen. Similar preparations from adult guinea pigs were active, 

 but those obtained from embryonic specimens were active only if a member 

 of the Kreb's cycle, such as fumarate, was added to the medium. Homogen- 

 ized rat liver required a pteridine derivative in addition to fumarate. 

 Surprisingly, 10-formylfolic acid, aminopterin, and A-methopterin were 

 almost as potent as folic acid. Attempts to replace methionine as the 

 methyl donor by choline, betaine, sarcosine, formate, and other com- 

 pounds were unsuccessful. Cohen prepared soluble enzyme systems by 

 centrifugation of homogenized rat and guinea pig liver and observed that 

 the creatine-forming activity of these preparations in the presence of GA, 

 ATP, methionine, and either oxygen or nitrogen was not enhanced by 

 additions of folic acid or of fumarate. It was evident, therefore, that the 

 latter were not required under these conditions for the methylation of GA. 

 Inasmuch as appropriate additions of mitochondria to the soluble enzyme 

 system restored the need of folic acid, fumarate, and oxygen, it appeared 

 that the maintenance of a high level of ATP in the presence of mitochondria 

 was dependent on an aerobic process involving folic acid or another pteri- 

 dine. 



Menne^^*' ^^* studied the formation of creatine in pulped muscle. Muscle 

 extracts were found to contain a myosin-like apoenzyme and a heat-stable, 

 water-soluble coenzyme. Atrophic and dystrophic muscles were unable to 

 synthesize creatine. Conditions for creatine synthesis in muscle were also 

 examined by Barrenscheen."^- ^*^ According to this author creatine is formed 

 in muscle by methylation directly from methionine and indirectly from 

 choline with trimethylamine oxide as an intermediate. The pherase acting 

 on methionine was cyanide-sensitive whereas the cholinepherase was cy- 

 anide-insensitive. Vignos and Cantoni^^'' prepared partially purified frac- 

 tions of a guanidoacetic acid methylpherase which were free of the enzyme 

 necessary for the synthesis of active methionine. 



'29 H. Borsook and J. W. Dubnoff, /. Biol. Chem. 171, 363 (1947). 

 '30 J. Vignos, Jr., and G. L. Cantoni, Federation Proc. 11, 399 (1952). 

 "1 S. Cohen, J. Biol. Chem. 193, 851 (1951). 



1" S. Cohen, Federation Proc. 11, 197 (1952); J. Biol. Chem. 201, 93 (1953). 

 1" F. Menne, Hoppe-Seyler's Z. physiol. Chem. 273, 269 (1942). 

 "4 F. Menne, Z. ges. exptl. Med. 112, 38 (1943). 



'3* H. K. Barrenscheen and J. Pany, Hoppe-Seyler's Z. physiol. Chem. 283, 78 (1948). 

 '3* H. K. Barrenscheen and M. Pantlitschko, Hoppe-Seyler's Z. physiol. Chem. 284, 

 250 (1949). 



