Studies of structure and physiological function of chymotrypsin inhibitors in hemolymph of Bombyx mori

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(1) Determination of amino acid sequence of chymotrypsin inhibitor (Fujii and Aso) :Basic chymotrypsin inhibitors, CI-1 and CI-b1 controlled by Ict-A complex gene were isolated from larval hemolymph of the silkworm, Bombyx mori according to the purification method established by us. Isolated CI-1 consisted of 62 amino acid and its molecular weight was calculated 7,065 on the basis of the amino acid sequence. Number of amino acid of CI-b1 was equal to the CI-1. (also consisted of 62 amino acids as well as CI-1). and a molecular weight of 7,065. Both CIs were sequenced amino acid. The amino acid sequences of CI-1 and CI-b1 were followed ; .CI-1 DKPTTEPICEQAFGNSGPCFAYIKLYSYNQKTKKCEEFIYGGCQGNCI-b1 -----K-----------K---DNRFITLAECEQKCIK----D-------Glu at the 6th amino acid of CI-1 was changed into Lys at CI-b1, Gln at the 44th into Lys, and Ile at the 51st into Asp.(2) Analysis of structure of chymotrypsin inhibitor, CI-8 gene (Fujii and Doira) :CI-8 cDNA was amplified by PCR with the fat body cDNA library using the synthesized oligonucleotides as probe based on both sequences of N-terminal and C-terminal amino acid sequences. CI-8 cDNA was cloned to pUC9 plasmid. and was sequenced nucleotide. It consisted of 1,185bp and the deduced amino acid sequence consisted of 395a.a. CI-8 cDNA contained the signal peptide consisted of 16 amino acids being rich with lyophilic amino acid such as Leu and Val.(3) Analysis of physiology of chymotrypsin inhibitors (Kawaguchi and Koga) :The radioisotope labeled CI-8 was injected into the 5th instar larvae. Abundant labeled CI-8 was detected in the fat body resident along epidermis and did not in the fat body near the midgut.