DANIEL I. ARNON 535 



viologcn dyes of lo^v redox potential or of producing Ho from re- 

 duced methyl viologen (82) . Both of these dark reactions are known 

 to be catalyzed by cell-free hydrogenases from heterotrophic anaerobes 

 (H4, 119). 



By contrast, the hydrogenase from Chromatium, in a cell-free state, 

 was found in a recent investigation to be capable of reducing viologen 

 dyes with H^, (115) and of evolving hydrogen in the dark from re- 

 duced methyl viologen (Table 9) . Experiments on the photoevolu- 

 tion of hydrogen from cell-free preparations of Chromatium are 

 currently vmder way in this laboratory. 



In other experiments with cell-free preparations of Chromatium, 

 we found that chromatophores contained succinic dehydrogenase. 



TABLE 9 



Hydrogen Evolution from Reduced Methyl Viologen by 



Cell-free Hydrogenase from Chromatium 



(Ogata, Nozaki, and Arnon, 115) 



/xmoles H2 evolved/10 min/mg chl. 



Complete system 12.8 



Methyl viologen omitted none 



Na2S204 omitted none 



Hydrogenase omitted none 



Complete system contained, in a final volume of 3.0 ml, cell-free suspension (PS) 

 containing 0.4 mg bacteriochlorophyll and 80 xmoles Tris buffer, pYi 7.2. 0.1 ml of 

 20*^^ KOH was present in the center well and 16 yumoles of methyl viologen were 

 added to the sidearm. Methyl viologen was reduced by adding Na2S204 to the same 

 sidearm while gassing with argon. The reaction was carried out at 30°C in the dark. 



As shown in Fig. 16, the addition of succinate reduced the Chromatium 

 cytochromes. The reduction by succinate was slow at first but be- 

 came accelerated with time (114). 



Since Chromatium cytochromes are oxidized by light (Fig. 7) and 

 reduced by succinate (Fig. 16) or thiosulfate (95) in the dark, it 

 seems permissible to regard succinate, thiosulfate, and other sub- 

 strates in bacterial photosynthesis as electron donors for the cy to- 

 chrome-chlorophyll system of the chromatophores. Following the 

 absorption of photons by chlorophyll the electrons donated by those 

 substrates are raised to a high reducing potential, at least equal to 

 that of molecular hydrogen. It follows from the electron flow theory, 

 that when these electrons do not return to the cytochrome-chlorophyll 

 complex by the cyclic route (Figs. 4 and 5) , but are instead trans- 

 ferred by hydrogenase to protons, that hydrogen gas will be produced. 

 A non-cyclic, light-dependent electron path of this sort, capable of 



