1766 



Bassham, Benson, Kay, Harris, Wilson and Calvin 



Vol. 76 



In other experiments" the Scenedesmus have been 

 kept at a steady state of light, temperature, CO2 

 pressure, etc., and constant C'K)2 specific activity 

 until successive samplings of the suspensions showed 

 uniform labeUng ("satiu-ation") of all the common 

 photosynthetic reservoirs (PGA, RDP, GMP, etc.). 

 The total CO2 pressure was then rapidly changed 

 from 1% C02-in-air to 0.003% in air, all other en- 

 vironmental conditions, including the specific ac- 

 tivity of C"02, being kept constant. The condi- 

 tions of this experiment were, therefore, similar to 

 those used previously' to study changing steady 

 state except that CO2 pressure was changed in- 

 stead of illumination. In the case where the CO2 

 pressure was lowered (Fig. 6), the initial effects on 

 the reservoir sizes of PGA and RDP were just the 

 opposite of those observed when the illumination 

 was stopped. Lowered COi pressure resulted in an 



RDP 



Triose phosphate 



4' 



B -«- 



A -*- 



20 



> 



a 

 Pi 



10 



1% CO, 



PGA 



RIBULOSE Old 

 AREA 



SCENEDESMUS 6' C 



-600 



-200 



-100 



45 minutes C"0, at 6° C. 

 Fig. 6. 



100 



Time in seconds. 



increase in the reservoir size of RDP and a decrease 

 in that of the PGA. After a time the reservoir of 

 RDP passed through a maximum and dropped to 

 a lower level but the new steady state RDP res- 

 ervoir was now greater relative to that of PGA. 

 The labeled glycolic acid present, though rather a 

 small percentage of total activity, increased many 

 fold when the COa pressure was lowered. The res- 

 ervoir of glycolic acid increased much more slowly 

 than that of the RDP and did not pass through a 

 corresponding maximum, thus eliminating the pos- 

 sibility that most of the labeled glycolic acid was 

 formed by thermal decomposition of RDP subse- 

 quent to killing of the cells. 



Discussion 

 1. Origin of PGA. — It has been suggested that 

 RDP is the compound which supplies the two- 

 carbon atoms for the carboxylation reaction lead- 

 ing to PGA.' If the reactions of these compounds 

 are represented by 



(19) A. T. Wilioo, Thesis, to be submitted as partial fulfillment ol 

 requirements for the degree of Doctor of Philosophy. UnlTenltjr of 

 California. 



Sugar rearrangements 



then the initial changes in reservoir sizes which 

 would accompany changes in light or COj pressure 

 can be predicted. When the light is tmned off, 

 reducing power [H] decreases, so the reservoir of 

 PGA would increase and that of RDP decrease. If 

 CO2 pressure decreases, then the reservoir of RDP 

 would increase and that of PGA would decrease. 

 Both effects, as well as those opposite effects which 

 would be expected to accompany a resumption of 

 light or increase in COj pressure, 

 have been observed. These re- 

 sults support the proposal of a 

 carboxylation of RDP to give 

 two molecules of PGA or the 

 reductive carboxylation to give 

 one molecule of PGA and one 

 of phosphoglyceraldehyde as 

 the first step in the path of 

 carbon dioxide reduction. 



It is also possible that the 

 products of this carboxylation 

 may be phosphoglyceraldehyde 

 and 3-phosphohydroxypyru- 

 vate. In this case subsequent 

 reduction of the phosphohy- 

 droxypyruvate would give first 

 PGA and then phosphoglycer- 

 aldehyde. The reaction of phos- 

 phoglyceraldehyde with hy- 

 droxypyruvate to give ribulose 

 monophosphate and COj has 

 been demonstrated by Racker^° 

 to take place under the influ- 

 ence of the transketolase en- 

 zyme. However, the increase in PGA concentra- 

 tion which is observed on stopping the illumina- 

 tion of photosynthesizing algae,' would probably 

 not be seen if a reduction of hydroxypyruvate 

 were required to form PGA since the reducing agent 

 would presumably no longer be formed in the dark. 

 Moreover, paper chromatographic analysis should 

 detect either phosphohydroxypyruvate or its de- 

 carboxylation product, phosphoglycolaldehyde, and 

 neither have been found in our experiments. When 

 C'*-labeled hydroxypyruvate was administered to 

 algae in this Laboratory, the labeled acid was me- 

 tabolized to give a variety of compounds, similar to 

 those formed from labeled pyruvate or acetate, which 

 were related more closely to the tricarboxylic acid 

 cycle and fat synthesis than to the compounds usu- 

 ally associated with carbon reduction in photosyn- 

 thesis. 



There remains the possibility that the RDP first 

 spUts to give a three-carbon molecule and a free 

 two-carbon fragment which is then carboxylated. 



(20) B. Racker, G. de la Haba and I. G. Leder, This Joijknal, Ti. 

 lOlO (1068). 



0003% COi 



200 



300 



98 



