130 BRITTON CHANCE 



that we observed at wavelengths appropriate to cytochrome c* and 

 omitted the quinohne oxide. Thus Fig. 14 represents an experiment in 

 which pigeon-heart mitochondria are pretreated with 4 niM succinate and 

 sufficient hydrosulphide to block respiration. Under these conditions 

 cytochrome c is completely reduced in about i min. As explained above, 

 pyridine nucleotide is not reduced. Upon addition of 530 /^M ATP, 

 pyridine-nucleotide reduction in agreement with Fig. 13 is observed. At 

 the same time we recorded an abrupt upward deflection of the spectro- 

 photometric trace indicating an oxidation of cytochrome c to an extent of 

 71% of the total. While pyridine-nucleotide reduction proceeds exponen- 

 tially toward a highly reduced level, the response of cytochrome c to ATP 



Cytochrome-c oxidation t 

 550-540mAt 



l-2//.M/sec 



Aerobic P. H. M 



+ 

 4mM succinate 

 360mM N-agS 



t I 



530 /iM ATP 



50 sec 



log Iq/I -0005 



Pyridine 

 nucleotide reduction , 

 365-450 _L ^ 



f^M 3-6mM DPNH 



Fig. 14. Cytochrome c oxidation and pyridine-nucleotide reduction in 

 succinate- and sulphide-treated mitochondria (concentrations indicated on figure). 

 Pigeon-heart mitochondria suspended in mannitol-sucrose-" tris " medium, pH 

 7-4, temperature 26, protein concentration i-i mg/ml. (Expt. i85b-3). 



addition is cyclic and reduction toward the initial level proceeds rapidly. 

 Other experiments show cytochrome a to be oxidized as well, cyto- 

 chrome b showing little initial change and then a slow reduction* which 

 is complete at about the same time as that of pyridine nucleotide. Studies 



* Lundegardh [22] has reported an effect of ATP (and fumarate) upon the 

 interaction of cytochromes b and ''dJi" in anaerobic wheat roots, cytochrome 

 "rf/i" being "turned over into a state of strong reduction under the influence of 

 ATP (and fumarate) simultaneously with cytochrome b remaining more oxidized". 

 The interpretation of this interesting result is rendered ambiguous by the fact 

 that the existence of cytochrome " dh" in wheat roots and other plant or animal 

 tissues has not been confirmed by other workers [23] nor by us using liquid nitrogen 

 spectroscopy of wheat-root mitochondria (B. Chance and W. Bonner, Jr., un- 

 published observations). 



