260 A SYMPOSIUM ON RESIRPATORY ENZYMES 



move the four-carbon acids more effectively. But this preparation 

 did not function in all respects like normal cells, failing to oxidize 

 glucose and succinate among others, although it did oxidize the four- 

 carbon acids (fumaric and malic) and lactate and pyruvate to acetic 

 acid. Since succinate was not oxidized, we believed that the cyto- 

 chrome system might have been injured, inasmuch as the hydrogen, 

 according to Szent-Gyorgyi, is transmitted through succinate to the 

 cytochrome system and thence to oxygen. When methylene blue or 

 cresyl blue was added to the system as carriers of hydrogen to re- 

 place the cytochrome system, no oxidation of succinate occurred. 

 Injury to Havoprotein action appeared ruled out because of the rapid 

 turnover of the substrates that were oxidized, i.e., fumarate, malate, 

 etc. Since acetone is known to destroy cytochrome oxidase, it was 

 difficult to understand the rapid attack on fumarate, malate, lactate, 

 and pyruvate. It was shown spectroscopically that the preparation 

 contained a cytochrome oxidase resistant to the acetone treatment. 

 These results were enough to indicate that investigations of bacterial 

 respiration were to prove interesting as well as a bit troublesome. 



One of our principal objectives has been to integrate the hetero- 

 trophic assimilation of carbon dioxide and respiration. We have 

 therefore run a number of experiments to gain a better insight into 

 the mechanism of bacterial respiration. The evidence so far ac- 

 cumulated seems to indicate that the Szent-Gyorgyi cycle does not 

 function in bacterial respiration. Glucose, for instance, is not at- 

 tacked by Micrococcus lysodeikticus anaerobically by cell suspen- 

 sions or the acetone preparation. If the Szent-Gyorgyi system were 

 operating, malic acid present would be oxidized anaerobically, since 

 fumarase would provide fumaric acid as a hydrogen acceptor for the 

 system. This does not occur. 



Thus far the evidence favors the occurrence in principle of the 

 Krebs citric acid cycle in bacteria. As the Krebs cycle is presented, 

 every alpha-keto acid with the single exception of pyruvic acid is 

 oxidatively decarboxylated. This point should be further investigated 

 to determine whether pyruvic acid is not also oxidatively decar- 

 boxylated, not necessarily to acetic acid, but to some two-carbon 

 compound which is able to condense with oxalacetic acid to initiate 

 the citric acid cycle. Acetic acid is commonly found as an end-prod- 

 uct when juices or perhaps injured cells are employed. Maintenance 

 of an adequate supply of oxalacetic acid is a requirement of the 

 Krebs scheme, and this is assured by regeneration in the cycle and 

 by the utilization of carbon dioxide through the Wood and Werk- 



