358 UNITY AND DIVERSITY IN BIOCHEMISTRY 



the separate operations of the pentose cycle, and, according to Calvin 

 (Fig. 94), of the associated photosynthetic cycle, have been reproduced in 

 vitro using purified enzymes. 



Aldolase catalyses the formation of heptulose diphosphate from a 

 molecule of tetrose phosphate and a molecule of triose phosphate 

 (C4 + C3 = C7). Phosphatase converts the heptulose-PP to heptulose-P 

 which, with a further molecule of triose-P, in the presence of transketolase, 

 forms a molecule of ribose-P and a molecule of ribulose-P (C7 + C3 = 2C5). 

 The isomerase for pentose phosphates converts this mixture to ribulose-P 

 which, in the presence of ATP and phosphopentokinase, gives ribulose-PP. 



However it still remains, in this enzymatic scheme, to explain the 

 fixation of CO2 when entering this series of reactions, Calvin has isolated 

 from spinach leaves and from ultrasonic macerates of the green alga 

 Chlorella a soluble enzyme which, in the presence of bicarbonate, converts 

 Ru-PP to two molecules of PGA. He has called it carboxydismutase, because 

 the carboxylation depends on the oxidation of C-3 of ribulose to a carboxyl 

 group. 



The fact that all the necessary enzymatic tools have been isolated does 

 not mean that the cycle actually does function. In fact for the reduction of 

 PGA to a triose, a molecule of ATP and a molecule of TPNH are required. 

 When we get to Ru-P, a further molecule of ATP is necessary for the 

 formation of Ru-PP. 



The cycle has been completely reconstructed in vitro by Racker (1955), 

 who has thus achieved the reductive synthesis of a sugar from CO2 and 

 plant enzyme extracts. The sequence of reactions is as follows: 



(1) 3 pentose-P + 3 ATP > 3 Ru-PP + 3 ADP 



(2) 3 Ru-PP + 3 CO2 + 3 H2O > 6 PGA 



(3) 6 PGA + 6 ATP > 6 DPGA + 6 ADP 



(4) 6 DPGA + 6 DPNH + 6 H+ > 6 triose-P + 6 DPN + 6P 



(5) 4 triose-P > 2 HPP 



(6) 2 HDP + 2 H2O > 2 HMP + 2 P 



(7) 1 HMP + 1 triose-P > 1 pentose-P + 1 tetrose-P 



(8) 1 HMP + 1 tetrose-P > 1 heptulose-P + 1 triose-P 



(9) 1 heptulose-P + 1 triose-P > 2 pentose-P 



sum (1-9) 3 CO2 + 9 ATP + 5 HoO + 6 DPNH + 6H+ 



> 1 triose-P + 9 ADP + 6 DPN -f 8 P 



(10) 9 H2O + 9 DPN > 9 DPNH + 9 H+ + 9 O 



(11) 3 DPNH + 9 ADP + 9P + 3H+ + 30 > 3 DPN + 



9 ATP + 12 H2O 



sum (1-11) 3 CO2 + 2 H2O + P > 1 triose-P + 6 O 



If we ignore equation (10) and (11) for the moment, we see that overall 

 (reactions 1-9), to introduce a CO2 molecule into a triosephosphate, three 



