458 



J. A. BASSHAM, M. KIRK 



that this percentage is a lower limit based only on absolute measurements of identified 

 compounds. 



A lower limit for the amount of carbon incorporated via C^ plus Cj carboxylation 

 is obtained by plotting Tg — ^Tjf (Fig. 7). The minimum rate of this incorporation is 

 about 0.4 /imoles/min/ml algae, or about 3 % of the total. Note that this value is 



I 



to 20 30 ■W 50 60 



Time in Seconds with ^^C02 



Fig. 7. Appearance of "C in compounds derived from PGA and in compounds derived from C, + Cj 

 carboxylation in Chlorella pyrenoidosa vs. time of photosynthesis with "COj. 



for the actual introduction of COj and does not include the carbon derived from PGA 

 (Tgf). The rate of incorporation of '*C into these three compounds thus accounts for 

 about 4 times 3, or 12 % of the total in this experiment. Other experiments indicate 

 that the relative contribution of C3-C1 carboxylation varies considerably and tends 

 to be higher (up to 3 times that reported in this case) when the rate of COg fixation is 

 greater and when amino acid synthesis is more rapid. In addition to the three com- 

 pounds listed here, other substances may be derived in part from C^-Cg carboxylation, 

 such as glutamic acid and citric acid, discussed below. 



While at least 73 % of the total rate of fixation of carbon has thus been shown 

 to be due to the carbon reduction cycle and C1-C3 addition, there is no indication 

 of any other significant fixation pathway. In Fig. 8 the ^*C found in glutamic acid 

 and in citric acid is shown. Could this labeling of glutamic acid be the result of a 

 carboxylation of y-aminobutyric acid? The maximum rate of labeling of glutamic 

 acid and in citric acid is shown. The maximum rate of labeling of glutamic acid is 

 about 0.7 /^moles/min or 4.5 % of all ^*C fixed. Since this rate is found between 5 and 

 20 min, it probably represents labeling of all five carbon atoms of glutamic acid, 

 because the precursors are surely at least partially labeled after 5 min. The labeling 

 due to carboxylation reaction would be expected to begin during the first 30 sec, if 

 one is to judge by the other known carboxylation reactions which were discussed 

 earlier. Yet, after the first 31.5 sec, the glutamic acid contains only 0.02 fimoles of 

 i*C. Between 40 and 60 sec, its labeling rate is only 0.2 /xmoles/min. Moreover, 

 y-aminobutyric acid itself would have to be synthesized from CO2 (by some as yet 



114 



