448 J- A. BASSHAM, M. KIRK 



INTRODUCTION 



Much of the biochemical pathway through which carbon dioxide is reduced during 

 photosynthesis in algae has been established^-^ A principal feature of this pathway 

 is the carbon reduction cycle. A simplified version of this cycle is given in Fig. i, 

 which shows the key steps. 



To map these paths, Calvin et al.^^" gave radioactive compounds, such as 

 "CO2 and KHj^^po^, to photosynthesizing plants. The plants made various reduced 

 organic compounds from these labeled substrates. They were then killed and the 

 soluble compounds were extracted from the plant material and analyzed by two- 

 dimensional paper chromatography and radioautography. The compounds were 

 identified and their radioactive content determined. From the amount and location 

 of radioactive elements within compounds following exposures of the plants for 

 various lengths of time and under various environmental conditions, biochemical 

 pathways were followed. 



Fig. I. Carbon reduction cycle (simplified version), (i) Ribulose diphosphate reacts with COj to 

 01-.OL1GO-. AND GLYCEROL PHOSPHATES givc an unstable six carbon compound which 



poLrssccHABiDES GALACTOSE PHOSPHATES spUts to give two three carbon compounds. At 



least one of these is 3-phosphoglyceric acid. The 

 other three carbon compounds might be either 

 ' ^'^^ 3-PGA, as it is known to be in the isolated en- 



PENTOSE-5-PHospHATES fHEPTOSE PHOSPHATES zyme System, or some other three carbon com- 



4TP (4 <HExosE PHOSPHATES pound such as a triose phosphate (dashed arrow) . 



\| JTRiosE PHOSPHATES ^^j pQ^ jg reduced to triose phosphate with 



RIBULOSE DIPHOSPHATE _^ ^^/ / j^jp ^^^ TPNH derived from the light reaction 



.^^TPNH 2^OTP^ ^^^ water. (3) Various condensations and re- 



arrangements convert the triose phosphates to 

 pentose phosphates. (4) Pentose phosphate is 



c .„„„j£ phosphorylated with.\TP to give ribulose di- 



-ALANINE phosphate. Further carbon reduction occurs via 



conversion of PGA to phosphoenolpyruvic acid, 

 (s) andcarboxylation, (61, to form a four carbon 

 compound (probably oxaloacetic acid). Keac- 

 -ASPARTicAcio tions leading to the formations of some of the 



secondary intermediates in carbon reduction are shown by the arrows lettered a through g. 



In the present study we have extended our information about these pathways 

 by more precise control of the environmental conditions during exposure of the 

 plants to tracers. At the same time we have made measurements of the rate of entry 

 of tracer into the plant and of the rate of appearance of the tracer in specific com- 

 pounds. 



We sought answers to the following questions: (a) How much of the total carbon 

 taken up by the plants enters the metabolic network via carboxylation of ribulose 

 diphosphate (reaction i)? (b) How much of the total carbon taken up enters by 

 carboxylation of PEPA (reaction 6)? (c) Are any other carboxylation reactions, such 

 as the carboxylation of y-aminobutyric acid", of any importance in steady state 

 photosynthesis? (d) Does the carboxylation of ribulose diphosphate in vivo lead to 

 one product only (PGA) or does it lead to two products (PGA and some other 3-carbon 

 compound)? 



"Steady state photosynthesis" as used in this paper, is defined as a condition 

 under which unicellular algae are carrying out the reaction of photosynthesis, are 

 synthesizing all of the normal cell constituents, and are growing and dividing at 



aTP =■ 



104 



