Oxygen-17 Appears Only in Protein in Water-Stressed Soybean Leaves Labeled by ¹⁷O₂

Abstract

We have used a rotational-echo adiabatic-passage double-resonance (13)C{(17)O} solid-state NMR experiment to prove that the glycine produced in the oxygenase reaction of ribulose bisphosphate carboxylase-oxygenase is incorporated exclusively into protein (or protein precursors) of intact, water-stressed soybean leaves exposed to (13)CO(2) and (17)O(2). The water stress increased stomatal resistance and decreased gas exchange so that the Calvin cycle in the leaf chloroplasts was no more than 35% (13)C isotopically enriched. Labeled O(2) levels were sufficient, however, to increase the (17)O isotopic concentration of oxygenase products 20-fold over the natural-abundance level of 0.04%. The observed direct incorporation of glycine into protein shows that water stress suppresses photorespiration in soybean leaves.

Carbon partitioning in soybean (Glycine max) leaves by combined 11C and 13C labeling

“…a, top left and right). This is the same conclusion that was reached earlier by using 17 O 2 labeling of the C 2 pathway in a water‐stressed whole soybean leaf with NMR detection of 17 O label only in protein, and not in starch or sugars (Gullion et al ., ).…”

“…Using solid‐state nuclear magnetic resonance (NMR), we have found that a fraction of the glycine from the photorespiratory C 2 cycle is not decarboxylated or returned to the Calvin cycle (Noctor et al ., ), but is inserted instead as glycyl residues in proteins (Cegelski & Schaefer, ; Gullion et al ., ). This result is consistent with earlier experiments using 14 CO 2 labeling and detection of label in both amino acids and proteins (Ongun & Stocking, ; Dickson & Larson, ).…”

Is the recovery of (photo) respiratory CO2 and intermediates minimal?

“…Other very similar studies taking advantage of 17 O– 13 C (Guillon et al ., ) or other hetero–nuclear interactions (Cegelski & Schaefer, ) and NMR technology have also suggested that under low CO 2 or water stress, glycine is not converted to phosphoglycerate and is instead incorporated into proteins. These studies are probably flawed as they have no consideration for 13 C enrichment in Ser C‐1 or C‐2; neglect 17 O/ 16 O exchange with solvent water (e.g.…”

RF field strength control and an envelope detector

“…In an extreme case, a 13 C-17 O REDOR experiment on soybean leaves required several months of NMR time. [3] In order to obtain quantitative results from NMR experiments taking a long time, the parameters of the experiment must be accurately controlled. Precise sample spinning speeds are necessary for rotor-synchronized pulse trains, [4] and most laboratories are equipped with good spinning speed controllers.…”

“…Magic angle sample spinning NMR experiments can last days and, possibly, weeks. In an extreme case, a 13 C– 17 O REDOR experiment on soybean leaves required several months of NMR time . In order to obtain quantitative results from NMR experiments taking a long time, the parameters of the experiment must be accurately controlled.…”