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Unambiguous assignment of the H3S and H3R deuterations of cerebral (2-13C) glutamate by 13C NMR at 18.8 tesla

✍ Scribed by Tiago B. Rodrigues; Inês R. Violante; Sebastián Cerdán

Publisher
John Wiley and Sons
Year
2010
Tongue
English
Weight
310 KB
Volume
63
Category
Article
ISSN
0740-3194

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✦ Synopsis

We used high-field 13 C NMR (18.8 T) to assign unambiguously the isotopic shifts induced by the deuterium substitutions of the H3 proR and H3 proS hydrogens of (2-13 C) glutamate in extracts of the brain from deuterated animals. Monodeuterated H3R or H3S glutamate diastereoisomers were produced stereospecifically either by chemical synthesis or by coupling the reactions of isocitrate dehydrogenase and aspartate aminotransferase in deuterated medium, respectively. We show that the (3S-2 H) or (3R-2 H) deuterations induce characteristic small (D 2 5 20.058 parts per million (ppm)) or large (D 2 5 20.071 ppm) vicinal isotopic shifts upfield of the perprotonated (2-13 C) glutamate resonance (at 55.5 ppm). Isotopically shifted (2-13 C, 3S-2 H) or (2-13 C, 3R-2 H) glutamate singlets are conveniently observed by high-field 13 C NMR in brain extracts from deuterated rats. Since the (3S-2 H) or (3R-2 H) glutamate diastereoisomers are produced stereospecifically by the cytosolic or mitochondrial isoforms of aconitase and isocitrate dehydrogenase, our results will facilitate the 13 C NMR investigation of these enzymatic activities and their role in subcellular glutamate trafficking.

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