Hydrochloric Acid → 2-dehydro-3-deoxy-D-gluconic acid

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Reaction Steps

Samples of the purified gluconate dehydratase solution prepared in Example 3, which was dissolved in the 30 mM Tris buffer (pH 8.5) containing 1 mM sodium D-gluconate and 1 mM magnesium chloride, were allowed to stand for 30 minutes, 2 hours, or 12 hours at 4, 20, 30, 40, 50, 55, 60, 65, or 70° C. Another sample was allowed to stand at 4° C. for 7 days and 30 days. Then, each sample subjected to heat treatment as above was reacted with 1 mL of a liquid containing 1 μmmol of Tris buffer (pH 8.5), 40 μmol of sodium D-gluconate, and 1 μmol of magnesium chloride at 37° C. After 10 minutes, 200 μL of 1 M hydrochloric acid solution was added to the reaction mixture to stop the reaction. Then, the amount of 2-keto-3-deoxy-D-gluconic acid produced was determined by high-performance liquid chromatography, and the reaction rate was calculated. The results are shown in Table 2 by relative activities to the enzyme activity before heat treatment which is represented as 100. The activity of 100% was maintained after treatment at 55° C. for 2 hours, and it is also stably maintained for 30 days at 4° C.

Provenance & Evidence