CYTOCHROMES 157 



transferring enzyme. The spectrum was obviously that of a hemin- 

 containing compound which resembled in type that of Spirographis 

 hemin (36). The alpha-band of the reduced carbon monoxide com- 

 plex lies at 5920 A. and the gamma-band at 4320 A. Finally, in the 

 highly respiring Bacterium Pasteurianum (Acetobacter pasteuria- 

 num), under anaerobic conditions, a weak band was observed at 

 5890 A. which was attributed by Warburg and Negelein (37) to the 

 reduced form of the oxygen-transferring enzyme itself, since carbon 

 monoxide shifted the band to 5920 A. On the other hand, cyanide 

 produced a band at 6390 A. which, since it may be observed even 

 in the simultaneous presence of the 5890 A. band, need not be a 

 derivative of the oxygen-transferring enzyme. Keilin (38) believes, 

 however, that the 5890 A. band is only a degradation product of 

 cytochrome a and is seen only in certain bacteria. In fact, Fujita 

 and Kodama (39) observed the 5890 A. band in bacteria only when 

 the cytochrome a band was absent and have named this band cyto- 

 chrome flj. Certainly the best criteria by which to establish the 

 identity of a compound with Warburg's oxygen-transferring enzyme 

 would be the positions of the carbon monoxide absorption bands 

 (5920 and 5320 A.). 



Keilin's work (40) with "indophenol oxidase" pointed to the 

 identity of this enzyme with the Warburg enzyme. The oxidase 

 brought about the aerobic oxidation of the cytochromes, was in- 

 hibited by cyanide, and showed a Hght-reversible inhibition with 

 carbon monoxide. 



Because of the similarity it has been generally believed that War- 

 burg's "oxygen-transmitting enzyme" and Keilin's presently-called 

 cytochrome oxidase are identical. Until recently, however, Keilin 

 had not observed a band that he could attribute to the carbon 

 monoxide complex of the oxidase. 



In 1939 Keihn and Hartree (10) believed that they had identified 

 spectroscopically in heart muscle extracts a new cytochrome, a^, 

 which might be identical with the oxidase. They concluded that 

 the cytochrome a band at 6000-6050 A. is actually due to com- 

 ponents a and a^, since upon addition of carbon monoxide this band 

 divides and a new one appears at 5900 A. With the aid of strong 

 cane sugar or glycerine solutions, or bile salts, to clarify the solutions 

 for spectroscopic examination, they were able to examine further 

 the Soret or gamma-bands of the cytochromes. Simultaneously with 

 the above shift, a portion of the 4480 A. band is shifted to 4320 A. 

 These two new bands represent the carbon monoxide complex of 



