450 



M.C\uviN ami 1'. Massini: Tlie TjIIi ot Carbon in I'hotosyulhesis 



[Exi'tKltNlIA\'0L.\'III/I2J 



The recent identification' as uridine diphospho- 

 glucose (U.D.P.G.) of the spot which had been previ- 

 ously* called «the unknown glucose phosphate spot» 

 has lead to another suggestion as to the mode of for- 

 mation of sucrose. Glucose-labeled U.D.P.G. appears 

 very early in the sequence of compounds formed. Fur- 

 thermore, it has been possible to demonstrate the pres- 

 ence in the hexose monophosphate area of a sucrose 

 phosphate by using a carefully selected phosphatase, 

 containing no invertase, in the treatment of this entire 

 phosphate area'. We have suggested, therefore, that 

 U.D.P.G. may be involved in sucrose synthesis in a 

 manner similar to that of glucose-1-phosphate in the 

 numerous phosphorylase reactions, with the difference, 

 however, that the acceptor of the glucose moiety would 

 be some phosphate of fructose, thus producing a sucrose 

 phosphate. Recent work by Putnam and Hassid' gives 

 further support to the idea that only phosphorylated 

 derivatives of glucose and fructose are involved in 

 sucrose synthesis in higher plants. They found that in 

 sucrose synthesis, from labeled glucose in leaf punches, 

 no free fructose was formed, although the sucrose be- 

 comes equally labeled in both the glucose and fructose 

 portions. Conversely, when labeled fructose is used, no 

 free labeled glucose appears, while the sucrose is uni- 

 formly labeled in both moieties. 



It is possible that compounds of the U.D.P.G. type 

 could be concerned in the transformation of sugars and 

 the subsequent incorporation into polysaccharides. 

 Uridine diphosphate would thus serve as a carbon 

 carrier in the same way that pyridine nucleotides and 

 flavonucleotidcs are involved in hydrogen transfer ; the 

 adenylic acid system in phosphate transfer ; and coen- 

 zyme A in the transfer of acetyl groups. There is already 

 some evidence for the existence of other members of 

 the uridine diphosphate group from our own work, as 

 well as that of others*. 



We may now turn our attention from the fate of the 

 glyceric acid to the problem of its origin. An exami- 

 nation of Table I indicates quite clearly that the first 

 position in the glyceric acid to become labeled is the 

 carboxyl group. As time proceeds, the other two carbon 

 atoms in the glyceric acid acquire radioactivity and it 



' J. G. Ulchana.n €t at., in press. - J. G. Blchanan, J. A. Uass- 

 HAM, A. A. UtNSON, D. F. Bradley, M. Calvin, L. L. Dals, M. 

 Goodman, P. M. Hayls, V. H. Lynch, L. T. Norris, and A. T. 

 Wilson, Phosphorus Metabolism, \'ol. II (Johns Hopkins Press, 

 Baltimore, Maryland. 1952), in press. 



' S. Kawaclchi, a. a. Benson, N. Calvin, and P. M. Hayls, 

 J. Am. Chem. Sor. 71, AV7 (1052). 



' J. G. Buchanan, J. A. Bassham, A. A. HtNSON, V. V. Uradllv, 

 .M. Calvin, L. L. Dals, M. Goodman, P. M. Hayes, V. H. Lynch, 

 L. T. Norris, and A. T. Wilson, Phospliorus Metabotism, \'ol. II 

 (Johns Hopkins Press, Baltimore, Maryland, 1902), in press. - J. G. 

 Buchanan, in press. 



• E. W. Putnam, Thesis (University of California, Berkeley. l'J5'.>). 



* R. Caputto, L. F. Leloir, C. E. Cardini, and A. C. Paladim. 

 J. Biol. Chem. IS4, 333 (1950). - A. C. Paladini and L. F. Leloir, 

 Biochem. J. 51, 126 (1951). - J. T. Park, J. Biol. Chem. 1S4, 885 

 (1952). 



appears that they acquire it at equal rates, at least 

 within the present accuracy of the experiments. 



It thus appears that the most rapid reaction which 

 carbon dioxide can undergo at least at high light in- 

 tensities, is a condensation with a Cj fragment leading 

 directly to phosphoglyceric acid. An examination of the 

 chromatograms of a very short photosynthetic period 

 shows glycine and glycolic acid as the only two-carbon 

 compounds present. The distribution of radioactivity 

 among the carbon atoms of these two compounds is 

 always equal and the same and corresponds very well 

 with that in the alpha- and beta-carbon atoms of the 

 glyceric acid, as may be seen from Table I. This sug- 

 gests that glycolic acid either is in the direct line for 

 the formation of the Q carbon dioxide acceptor, or is 

 very closely related thereto. 



The question now arises as to the source of this Cj 

 carbon dioxide acceptor. There are, of course, only two 

 possibilities for its origin. Either it results from a one- 

 plus-one combination or it must result from the split- 

 ting of a four-carbon compound or a larger one. In order 

 for it to result from the combination of two one-carbon 

 fragments there must exist as an intermediate some 

 one-carbon compound more reduced than carbon diox- 

 ide which, in turn, may combine either with itself or 

 with carbon dioxide. Furthermore, the reservoir of 

 this one-carbon intermediate would have to be vanish- 

 ingly small since all attempts to find labeled, reduced, 

 one-carbon compounds, such as formic acid or formal- 

 dehyde, in the early stages of photosynthesis have 

 failed and, in addition, the resulting two-carbon frag- 

 ment is very nearly equally labeled in both carbon atoms. 

 One would also expect that these one-carbon com- 

 pounds would tend to disappear under conditions of 

 low carbon dioxide concentrations leading to the disap- 

 pearance of the two-carbon condensation product re- 

 sulting from them. This leads us to the supposition that 

 the formation of glycolic acid would be expected to 

 drop off under conditions of low carbon dioxide con- 

 centration which is the reverse of what is observed. 



We are thus left with the following possibility for the 

 C'a compound -the cleavage of some C4 or larger struc- 

 ture. The fact of the early appearance of label in malic 

 acid, taken together with the lack of any appreciable 

 amounts of label in the compounds of the tricarboxylic 

 acid cycle', led us to the supposition that malic acid 

 was either a precursor to, or very closely related to, a 

 four-carbon compound which could be split to produce 

 the required two-carbon fragment. 



In the course of the search for the two-carbon ac- 

 ceptor, and its immediate precursors, two new com- 

 pounds were identified as early products of carbon 

 dioxide incorporation which seem to have little to do 

 with the direct synthesis of hexoses and, therefore, had 

 a very likely function in the regeneration of the two- 



1 A. A. Benson and M. Calvin, J. Exptl. Botany ;, 63 (1950). 



84 



