TDP-4-keto-6-deoxyglucose

For more detailed studies of the overall pathway, a separation of the individual enzymes was carried out. For this purpose a single general procedure, effective in measuring 3,5-epimerase activities, was developed incubation of TDP-4-keto-6-deoxyglucose-3T (35, 36) in the presence of the enzyme resulted in tritium exchange with the medium. After completion of the incubation, further tritium exchange with the medium was prevented by the addition of sodium borohydride (Figure 7). 

Only in the presence of 3,5-epimerase could a significant exchange of tritium be observed. One line of evidence consistent with this interpretation came from the use of a mutant strain E. coli Y-10, incapable of producing 6-deoxyhexose. When extracts of E. coli Y-10 were incubated with TDP-4-keto-6-deoxyglucose-3T, no exchange with the medium was observed. 

GERK Streptomyces sp. KCTC 0041BP TDP-4-keto-6-deoxyglucose reductase... 

The same conclusion was drawn by Nelsestuen and Kirkwood (1971) who investigated the mechanism of the E. coli enzyme with UDP-4-keto-6-deoxyglucose and TDP-4-keto-6-deoxyglucose as substrates. The enzyme was reduced with NaB H4, yielding a product which contained tritium-labeled DPNH. Addition of the substrate led to rapid oxidation of the epimerase and to the formation of nucleotide sugars, from which fu-cose and quinovose were isolated after acid hydrolysis. Degradation of the sugars showed that the label was present exclusively in position 4. 

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