Molecular Plant Physiology and Environmental Biology, Research School of Biological Sciences, The Australian National University, Canberra, Australian Capital Territory, Australia.
The carboxylation kinetic (stable carbon) isotope effect was measured for purified d-ribulose-1,5-bisphosphate carboxylases/oxygenases (Rubiscos) with aqueous CO(2) as substrate by monitoring Rayleigh fractionation using membrane inlet mass spectrometry. This resulted in discriminations (Delta) of 27.4 +/- 0.9 per thousand for wild-type tobacco Rubisco, 22.2 +/- 2.1 per thousand for Rhodospirillum rubrum Rubisco, and 11.2 +/- 1.6 per thousand for a large subunit mutant of tobacco Rubisco in which Leu(335) is mutated to valine (L335V). These Delta values are consistent with the photosynthetic discrimination determined for wild-type tobacco and transplastomic tobacco lines that exclusively produce R. rubrum or L335V Rubisco. The Delta values are indicative of the potential evolutionary variability of Delta values for a range of Rubiscos from different species: Form I Rubisco from higher plants; prokaryotic Rubiscos, including Form II; and the L335V mutant. We explore the implications of these Delta values for the Rubisco catalytic mechanism and suggest that Rubiscos that are associated with a lower Delta value have a less product-like carboxylation transition state and/or allow a decarboxylation step that evolution has excluded in higher plants.