THE CHEMISTRY OF PHOTOSYNTHESIS 133 



tion of enzymes or cofactors to the suspension. The rates of both reactions 

 were about the same as those observed in intact cells. These findings are in 

 contrast to those of Arnon, who stated that the addition of certain enzymes 

 and cofactors is indispensable for obtaining" photosynthetic phosphorylation 

 and CO2 uptake in isolated chloroplasts and chloroplast fragments. Thomas 

 et al. explain the discrepancy as follows. Arnon used spinach chloroplasts 

 which are of the granulated type, whereas Spirogyra chloroplasts are lamellated, 

 spiral-shaped chloroplasts that are grana-free. In the latter thin layers of 

 stroma occur between bundles of lamellae (see § 3). This layered structure 

 may be capable of protecting the stroma against damage far more effectively 

 than may be possible in granulated chloroplasts. The lamellae are highly 

 resistant to experimental conditions (crushing of the cells), whereas the granu- 

 lated chloroplasts are easily damaged. The presence of intact stroma seems 

 to be necessary if complete photosynthesis is to be obtained. Either the 

 stroma contains important enzymes and cofactors or it protects enzymes lo- 

 cated on the surface of the lamellae. 



According to Arnon, the light reaction in photosynthesis only serves for 

 the production of ATP and TPNH, these substances being necessary for the 

 dark CO2 fixation reaction : 



dark 

 CO2 + 2 TPNH + 2 H+ + 2 ATP ^ (CH.O) + H2O + 2 ADP + 2 ph 



Arnon (5, 8, 56) found that the assimilatory power, i.e., ATP and TPNH, 

 generated in the light in the absence of COo in the green water-insoluble parts 

 of the chloroplasts (grana) is able to carry out CO2 fixation in the dark in 

 chlorophyll-free extracts (stroma) obtained by suitable centrifugation of the 

 chloroplasts. These extracts contain the assimilatory power produced in 

 the previous light reaction together with soluble enzymes. Small amounts of 

 glucose- 1 -phosphate and ^^C02 were added. The dark fixation of '^COo 

 by the chlorophyll-free extract to which assimilatory power was added was 

 of the same order of magnitude as CO2 fixation in the light in the presence of 

 ^^G02 since the commencement of illumination. The products of CO2 fixa- 

 tion were identical in both cases, as was shown by paper chromatography and 

 autoradiography. Instead of using ATP and TPNH generated by a light 

 reaction, both substances were supplied from external, chemical or enzymati- 

 cal sources. They were as effective in producing CO2 fixation as the assimi- 

 latory power generated by illuminated chloroplasts. 



Thus, Arnon found a physical separation of the light and the dark phase 

 through the identification of ATP and TPNH produced in the light and the 

 presence of CO2 fixing enzymes in aqueous extracts of chloroplasts. The 

 light phase is localized in the grana and the dark phase in the stroma (5, 8, 

 56). However, the necessity of the TPN+-reducing factor shows that, 

 in the light, some stroma factors must be involved. It follows from these im- 

 portant investigations that CO2 fixation of isolated chloroplasts must be con- 

 sidered to be a process independent of the photosynthetic pigment system. 



