ORIENTATION-CONSERVED TRANSFER OF SYMMETRICAL KREBS CYCLE INTERMEDIATES IN MAMMALIAN TISSUE

Metabolism of [2-C-13]-, [3-C-13]-, and [1,2,3-C-13] propionate in per fused rat livers and [2-C-13]acetate in perfused rat hearts has been e xamined in tissue extracts by C-13 NMR. Label from [2-C-13]propionate was preferentially incorporated into the C2 carbon of lactate, alanine , and aspartate in liver tissue while label from [3-C-13] propionate a ppeared preferentially in the C3 carbon of those same molecules. These data suggest that C-13 may not be completely randomized in the symmet ric citric acid cycle intermediates succinate and fumarate as is norma lly assumed but that some fraction of those intermediates may be trans ferred between enzymes in this span of the cycle with conservation of spatial orientation, consistent with recent results obtained in yeast [Sumegi et al. (1990) Biochemistry 29, 9106-9110]. This was confirmed by performing similar experiments with [1,2,3-C-13] propionate. Time-d ependent asymmetry was also observed between the intensities of the gl utamate C2 and C3 resonances and between the aspartate C2 and C3 reson ances in C-13 NMR spectra of intact hearts and heart extracts during e arly perfusion with [2-C-13]acetate. A model is presented which predic ts that isotopic asymmetry is observed only during the first 2-3 turns of the cycle pools when isotope enters the cycle via acetyl-CoA even if all symmetric cycle intermediates retain a unique molecular orienta tion on each pass through the citric acid cycle.