Pyruvic acid phosphate, labeled with

Our laboratory has studied the stereochemistry of methyl group formation in a number of a, 0 elimination reactions of amino acids catalyzed by pyridoxal phosphate enzymes. The reactions include the conversions of L-serine to pyruvate with tryptophan synthase 02 protein (78) and tryptophanase (79), of L-serine and l-tyrosine with tyrosine phenol-lyase (80), and l-cystine with S-alkylcysteine lyase (81). In the latter study, the stereospecific isotopically labeled L-cystines were obtained enzymatically by incubation of L-serines appropriately labeled in the 3-position with the enzyme O-acetyl serine sulfhy-drase (82). The serines tritiated in the 3-position were prepared enzymatically starting from [l-3H]glucose and [l-3H]mannose by a sequence of reactions of known stereochemistry (81). The cysteines were then incubated with 5-alkyl-cysteine lyase in 2H20 as outlined in Scheme 19. The pyruvate was trapped as lactate, which was oxidized with K2Cr202 to acetate for analysis. Similarly, Cheung and Walsh (71) examined the conversion of D-serine to pyruvate with... 

Fig. 4. Carbon-13 labelling routes revealing the principle of C4-C3 backflux identification by NMR upon incubation of C. glutamicum with [l- C]glucose. Considering that C-2 of the triose phosphates is unlabelled (to first approximation), the C balance for pyruvate C-2 reads P2 x (G -i- Adc) = (G x 0 -i- A c x 02), with G the glycolytic flux at the level of the triose phosphates, A c the C4-decarboxylating flux, and P2 and 02 the positional C enrichments of pyruvate C-2 and oxaloacetate C-2, respectively. Thus, the flux ratio Adc/(G + Adc) is equal to the measured ratio P2/02. Analogously, subtracting the C balances for oxaloacetate C-2 and C-3 yield (03-02) X (T -I- Acx) = (P3-P2) x Acx, with T the activity of the citric acid cycle, Acx the C3-carboxylating flux, and P3 and 03 the positional C enrichments of pyruvate C-3 and oxaloacetate C-3, respectively. Thus, the flux ratio A x/fT + A x) is equal to the measured ratio (03-02)/(P3-P2)...

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