414 Discussion 



4. Sulphite, or its biochemical equivalent, is the one established inter- 

 mediate in sulphate reduction. Since there is some reason to believe that 

 Cg is concerned in the functioning of the sulpliite reductase, it follows that 

 it must equally be involved in the reduction of sulphate. 



It is reasonable to conclude that cytochrome c^ is the sole haematin com- 

 pound among several electron transport enzymes of a conventional character 

 linking sulphate reduction to dissimilation processes in D. desulphuricans. 

 In aerobic organisms, cytochrome c^ is replaced by the cytochromes of the 

 a, b and c types (or an analogue of this system) and the function of the 

 sulphate reductase is taken over by cytochrome oxidase. 



Since this contribution was prepared, Ishimoto and colleagues have 

 obtained a cell-free sulphate reductase which requires adenosine triphosphate 

 (ATP). The reduction of sulphate by this preparation is stimulated by 

 addition of cytochrome c^. These experiments are mentioned by Egami, 

 Ishimoto and Taniguchi in their contribution to this pubhcation (see p. 404). 



REFERENCES 

 Ishimoto, M., Kondo, Y., Kamayama, T., Yagi, T. & Shiraki, M. (1958). Proc. int. 



Symp. Enzyme Chem. {Tokyo) p. 229. 

 Moses, V. (1955). J. gen. Microbiol. 13, 235. 

 PoSTGATE, J. (1956). J. gen. Microbiol. 14, 545. 

 PosTGATE, J. (1958). Prod. Mon. 22, 12. 

 PoSTGATE, J. (1959). Annu. Rev. Microbiol. 13, 505. 

 Senez, J. C. & PiCHiNOTY, F. (1958). Biochim. biophys. Acta 28, 355. 



DISCUSSION 



Nature and Properties of Cytochrome Cg 



Crystallization of cytochrome C3 



HoRio: Kamen and I, very recently, succeeded in crystallization of cytochrome Cg from 

 dried cells of Desulphovibrio desulphuricans which had been kindly sent by Postgate. 



To our regret, the general properties have not yet been studied. 



However, the story about the native and modified forms is the same in the case of 

 cytochrome C3 as it is with typical cytochrome c's. 



As indicated by Postgate, this bacterial cytochrome is the first one extracted from 

 bacteria which has a high iso-electric point. This property makes its purification on 

 Amberlite CG-50 quite effective, unlike the Pseudomonas cytochromes. 



With the lyophilized cells, most of cytochrome C3 is easily extractable with the use 

 of 01 M sodium citrate at a neutral pH, and it can be chromatographed with the use 

 of 015m (as NH4+) ammonium phosphate buff"er (pH 70). In addition to that 

 fraction which is not adsorbed on the resin, cytochrome c^ is chromatographed into 

 4 fractions. One of the fractions is eluted with 005 n ammonia after the chromato- 

 graphy with the buffer. 



From one of the fractions, cytochrome c^ has been crystallized. The other fractions 

 have been found to be artificially made by treating the crystalline sample under drastic 

 conditions; high and low pH, and boiling. Therefore, we can say that the crystallized 

 sample is most plausibly native, and that cytochrome C3 is fairly unstable to these 

 treatments. 



