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Retention of the Oxygen Atoms at Carbon-2 and Carbon-3 during the Carboxylation of Ribulose 1,5-bisphosphate

1978; Wiley; Volume: 89; Issue: 1 Linguagem: Inglês

10.1111/j.1432-1033.1978.tb20894.x

1432-1033

George H. Lorimer,

Microbial metabolism and enzyme function

Ribulose-1,5-bisphosphate carboxylase was allowed to react with [2-18O]ribulose 1,5-bisphosphate in the presence of either (a) carbon[18O]dioxide and [18O]water, or (b) unlabeled carbon dioxide and unlabeled water. The resultant 3-phosphoglycerate was isotopically analysed as trimethylsilyl glycerate by combined gas chromatography/mass spectrometry. In the former case the mole fractions of unlabeled, singly labeled, doubly labeled and triply labeled 3-phosphoglycerate were in the proportion: where n is the atom fraction 18O present in the water, in the carbon dioxide and in the 2-keto position of ribulose 1,5-bisphosphate. This result indicates that the oxygen atom at carbon-3 of ribulose 1,5-bisphosphate is retained during the carboxylation reaction to become one of the two carboxyl oxygen atoms of the 3-phosphoglycerate derived from carbon atoms 3–5 of ribulose 1,5-bisphosphate. In the latter case, significant (about 33% of maximum) quantities of 18O were found attached to carbon-2 of 3-phosphoglycerate, indicating that the 2-keto oxygen atom of ribulose 1,5-bisphosphate is retained during the carboxylation reaction to become the oxygen atom at carbon-2 of the 3-phosphoglycerate contining the newly incorporated carbon dioxide. The failure to observe maximum retention of the 18O is ascribed to the rapid exchange of the 2-keto 18O atoms of ribulose 1,5-bisphosphate with the 16O atoms of water. The results set constraints upon the mechanisms of carboxylation which can be accomodated, excluding those which would result in the loss of the oxygen atoms at either carbon-2 or carbon-3 of ribulose 1,5-bisphosphate.