| Budget Amount *help |
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1996: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1995: ¥1,400,000 (Direct Cost: ¥1,400,000)
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| Research Abstract |
Initiatorin, an endopeptidase localized in the glandula (g.) prostatica of the silkmoth, Bombyx mori, triggers the arginine degradation cascade, a specific energy-yielding system for spermatozoa. This protease activates the precursor of arginine carboxy-peptidase which acts at the second step of the arginine degradation cascade by producing free arginine from the Arg-peptide splitted by initiatorin from a substrate protein. This multi-functional enzyme also initiates the dissociation of eusperm bundles and motility of apyrene sperm. The secreting feature from cells of the g. prostatica to its lumen was immunohistologically observed with antiserum which had been prepared from a purified initiatorin preparation. Initiatorin was isolated from the two organs, the g. prostatica and spermatophore, separately. In the former purification procedures hydrophobic interaction chromatography and cation exchange chromatography were applied, whereas affinity chromatography on p-aminobenz- amidine was
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in the latter. These obtained initiatorins were assigned by their N_<alpha>-bezoylarginine ethyl ester-hydrolysing activity and also immunoblot after development of SDS-acrylamide gel-electrophoresis. Two types of initiatorins of molecular weights of 29 and 26kDa were isolated from the spermatophore, while three types of 30, 29 and 26kDa were from the g. prostatica. Ten N-terminal amino acid sequence of these initiatorins were identical. Prostatic 30kDa-initiatorin showed the strong induction of apyrene motility, but spermatophore 29kDa initiatorin did weak induction. 30kDa initiatorin satisfied the other three conditions as the multi-functional enzyme. Considered with highly strong self-degradation of initiatorin, 30kDa initiatorin produced in the cells of the g. prostatica must degrade to other initiatorins with smaller molecular weights in the g. prostatica and also in the sprmatophore. The initiatorin inhibitor localized in the g. prostatica seems to have an effect to protect cells to avoid the attack from this drastic protease. After unbinded from this inhibitor on ejaculation, initiatorin apparently "activated" causes many various reactions for sperm maturation. Fertilized eggs were laid, when a mixture of initiatorin and the content of the vesicula seminalis containing eusperm bundles and apyrene sperm was injected to the bursa copulatrix of virgin female moths instead of the spermatophore. Therefore, initiatorin is the indispensable factor for sperm maturation which causes the exact fertilization and fecundity. Less
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