The Biosynthesis of Porphyrins 249 



conclusions can be tested experimentally by blocking reaction F with 

 malonate. If these considerations are valid and if reaction C occurs, 

 the degree of labeling found in protoporphyrin from carboxyl-labeled 

 succinate should not be influenced by the presence of malonate. On 

 the other hand, methylene-labeled succinate can produce labeled por- 

 phyrin via reaction F. In this case the degree of labeling from 

 methylene-labeled succinate should be lowered in the presence of malo- 

 nate. It was found experimentally that the C 14 activity of the porphy- 

 rin synthesized from carboxyl-labeled succinate in the presence or 

 absence of malonate was the same, whereas that of the porphyrin 

 synthesized from methylene-labeled succinate was 50-6G"/o lower in 

 the presence of malonate than in its absence. 14 These experiments 

 demonstrated that the succinate can be converted to "active" succinate 

 via two pathways and that the malonate effect is a reflection of the 

 positions in the succinate which contain C 14 . 



It then became of interest to find the mechanism by which the 

 "active" succinate and glycine combine to form the pyrrole unit of 

 the porphyrin. It was realized that in the initial condensation of 

 glycine and succinate the whole molecule of glycine is involved, since 

 in all experiments in which glycine-2-C 14 was the substrate the carbon 

 atom in the pyrrole ring and the methene bridge carbon atom (Fig. 3) 

 had the same C 14 activity and no derivative of the a-carbon atom of 

 glycine (CH 3 OH, H 2 CO, HCOOH, CH 3 NH 2 ) could substitute for 

 glycine. These findings led us to the conclusion that the same deriva- 

 tive of glycine was utilized for the pyrrole ring carbon atom and for 

 the bridge carbon atom, even though the bridge carbon atom was no 

 longer attached to the nitrogen atom of glycine as is the ring atom. 

 On consideration of the possible method of condensation of succinate 

 and glycine, which would give rise to a product from which a pyrrole 

 could reasonably be made, the suggested mechanism must also account 

 for a method by which the carboxyl group of glycine is detached from 

 its a-carbon atom, subsequent to the initial condensation, for the 

 carboxyl group of glycine is not utilized for porphyrin synthesis. The 

 condensation of succinate on the a-carbon atom of glycine to form 

 a-amino-/?-ketoadipic acid (Fig. 7) would appear to be in agreement 

 with all the experimental findings and conclusions. The compound 

 formed, being a /?-keto acid, could then readily decarboxylate and thus 

 provide a mechanism by which the carboxyl group of glycine is de- 

 tached from its a-carbon atom subsequent to the initial condensation 

 of the whole molecule of glycine with succinate. Further, the product 



