CYCLIC AND NONCYCLIC PHOTO PHOSPHORYLATION 185 



The TPN- reducing factor in the water-soluble extract was found 

 to be heat stable and was identified as DPN by paper chromatography 

 after isolation withtheaidof aDuolite A-2 resin (28). Fig, 1 shows that 

 chromatophores photoreduced TPN in the presence of either catalytic 

 amounts of DPN or the boiled water-soluble extract of R. rubnim cells. 



The fact that TPN reduction was mediated by catalytic amounts of 

 DPN indicated that the chromatophores contained a transhydrogenase. 

 This conclusion is supported by the results shown in Fig. 2, Transhy- 

 drogenase activity in washed chromatophores was determined by meas- 

 uring the TPN reduced in the dark (as a change in optical density at 

 340 m^) in the presence of an added alcohol dehydrogenase system 

 and catalytic amounts of DPN. TPN was not reduced without DPN. 

 DPN was added either as the pure chemical or as a cell extract, i.e., 

 the boiled and Duolite A 2-treated supernatant solution from the chro- 

 matophore preparation. 



III. NONCYCLIC PHOTOPHOSPHORYLATION 



As already mentioned, we have interpreted the photo reduction of 

 DPN as evidence for a noncyclic electron flow in R. rubrum chroma- 

 tophores. To demonstrate noncyclic photophosphorylation in chroma- 

 tophores, it was necessary to establish that the photoreduction of 

 DPN was accompanied by ATP formation under conditions such that 

 ATP formation by cyclic photophosphorylation was excluded. 



Effect of antimycin A 



Photoreduction of DPN by succinate is inhibited by antimycin A (22). 

 However, antimycin A does not inhibit the photoreduction of DPN by 

 the ascorbate-DPIP couple (22). It should be noted that the photore- 

 duction of DPN by the ascorbate-DPIP couple in the presence of 

 antimycin A, which we reported previously (see Fig. 2 in ref. 22), oc- 

 curred at a concentration of ascorbate (6.7 x 10-3 m) at which cyclic 

 photophosphorylation catalyzed by DPIP is suppressed (Table 7), 

 Thus, the ATP formation (shown again in Table 8) which could not 

 have occurred via cyclic photophosphorylation because of the high 

 ascorbate concentration, could only have resulted from noncyclic 

 photophosphorylation. 



Requirement for electron donor and acceptor 



Table 8 shows ATP formation by chromatophores in the presence 

 of antimycin A, DPIP, and 6.7 x 10-3 m ascorbate. Appreciable photo- 

 phosphorylation occurred only in a complete noncyclic electron trans- 

 port system, i,e,, in the presence of both an electron donor (ascorbate 

 + DPIP) and an electron acceptor (DPN), Little photophosphorylation 

 occurred when either the electron donor or the electron acceptor, or 

 both, were omitted. Table 8 also shows that little photophosphorylation 



