| Budget Amount *help |
¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1992: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1991: ¥1,000,000 (Direct Cost: ¥1,000,000)
|
|---|
| Research Abstract |
To elucidate molecular mechanism of digestion and absorption in the silkworm, Bombyx mori, hydrolysis and absorption of sucrose was studied in the midgut, and in particular, sucrase in the midgut tissue was investigated.
Optimum conditions for assay of midgut sucrase in the fifth instar larvae and in pharate adults were different. Based on this result, optimum assay conditions for each developmental stage was selected. From the fourth instar until adult emergence, activity was determined. Maximum activity was obseved in the middle of the fifth instar. Electrophoretic pattern of soluble sucrase during development showed difference between larvae and pharate adults. The transition was at larval-pupal ecdysis. Ninety % of activity of midgut sucrase localizes in the cytosol and 10 % in the membrane-bound fraction in the fifth instar and pharate adult midgut. In vitro experiments using isolated midgut sac, it was demonstrated that membrane-bound sucrase is involved in hydrolysis of sucrose in the midgut and that peritrophic membrane enhances transport of hydrolysis products of sucrose accross midgut tissue into the haemocoel. Soluble sucrase was purified to near homogeneity using ammonium sulfate fractionation, and column chromatographies of ion exchange cellulose, hydroxylapatite, and Sephacryl as a chromatomedia. The enzyme was disclosed beta-fructofuranosidase from its kinetic properties. Solubilization procedure of membrane-bound sucrase with detergent was established and isolated sucrase showed properties distinct from soluble beta-fructofuranosidase.
|
