SEVERO OCHOA 



(20,21), it will be convenient to discuss it first because the methods 

 used in the study of this system permit a clearer picture of the pat- 

 tern into which the cellular mechanisms of carbon dioxide fixation 

 can be fitted. The primary fixation reaction involves the reversal of 

 the decarboxylation of oxalosuccinic acid to a-ketoglutaric acid and 

 carbon dioxide (reaction la). This reaction is catalyzed by an 



OXALOSUCCINIC CARBOXYLASE 



COOH COOH 



CO 



CO 



(la) 



enzyme, oxalosuccinic carboxylase, present in heart muscle and prob- 

 ably in other animal and plant tissues. The enzyme requires either 

 magnesium or manganese ions for activity. 



The equilibrium of reaction la is so far to the left that the avail- 

 able analytical methods would fail to show a formation of oxalosuccinic 

 acid even when starting with very high concentrations of a-ketoglutaric 

 acid and carbon dioxide in the presence of the enzyme. However, 

 reversibility can easily be demonstrated by adding isocitric dehydro- 

 genase (see Table I) and reduced triphosphopyridine nucleotide. 

 When this is done, the oxalosuccinic acid formed by carboxylation is 

 reduced to /-isocitric acid by TPNH2 which, in turn, is oxidized to 

 TPN (reaction lb): 



COOH 



I 

 CO 



ISOCITRIC DEHYDROGENASE 



COOH 



HC— COOH 



CH2 



I 

 COOH 



Oxalosuccinic acid 



+ TPNH2 



CHOH 



HC— COOH -f TPN 



I 

 CH2 



I 

 COOH 



/-Isocitric acid 



(lb) 



Reaction lb is the second step of the series by which carbon 

 dioxide is fixed in this system. The combined result of reactions la 

 and lb is reaction Ic. 



172 



