| 摘要 |
PURPOSE: To efficiently obtain the subject enzyme capable of accurately measuring fructose and useful for diagnosis, etc., of diabetes by culturing a microorganism, belonging to the genus Comamonas in a culture medium and capable of producing a sorbitol 6-phosphate dehydrogenase and collecting the resultant product from the cultured product. CONSTITUTION: A microorganism, belonging to the genus Comamonas and capable of producing a sorbitol 6-phosphate dehydrogenase [e.g. Comamonas sp. SY-77-1 (FERM BP-4810)] is inoculated into a culture medium, cultured at 28 deg.C for 20hr using a shaking culture machine at 120r.p.m. and the culture solution is then centrifuged to collect a microbial cell, which is subsequently suspended in a 20mM tris-hydrochloric acid buffer solution (pH8.5) and crushed with an ultrasonic crusher. The resultant crushed liquid is then centrifuged to collect a supernatant, which is subsequently passed through an anion exchange column to elute an adsorbed substance. The prepared eluate is then treated and purified by a hydrophobic column chromatography. Thereby, the objective sorbitol 6-phosphate dehydrogenase, having sufficient stability, capable of accurately determining fructose and used for monitoring, etc., of diabetes is efficiently obtained. |
