A fibroin secretion-deficient silkworm mutant, Nd-sD, provides an efficient system for producing recombinant proteins.

Full Abstract

The silkworm Nd-s(D) mutant is silk fibroin-secretion deficient. In the mutant, a disulfide linkage between the heavy (H) and light (L) chains, which is essential for the intracellular transport and secretion of fibroin, is not formed because of a partial deletion of the L-chain gene. To utilize the inactivity of the mutant L-chain, we investigated the possibility of using the Nd-s(D) mutant for the efficient production of recombinant proteins in the silkworm. A germ line transformation of the mutant with a normal L-chain-GFP fusion gene was performed. In the transgenic mutant, normal development of the posterior silk gland (PSG) was restored and it formed a normal cocoon. The biochemical analysis showed that the transgenic silkworms expressed the introduced gene in PSG cells, produced a large amount of the recombinant protein, secreted it into the PSG lumen, and used it to construct the cocoon. The molar ratio of silk proteins, H-chain:L-chain-GFP:fibrohexamerin, in the lumen and cocoon in the transgenic silkworm was 6:6:1, and the final product of the fusion gene formed about 10% of the cocoon silk. This indicates that the transgenic mutant silkworm possesses the capacity to produce and secrete the recombinant proteins in a molar ratio equal to that of the fibroin H-chain, contributing around half molecules of the total PSG silk proteins.

Author information

Author/s: Inoue, Satoshi (S); Kanda, Toshio (T); Imamura, Morikazu (M); Quan, Guo-Xing (GX); Kojima, Katsura (K); Tanaka, Hiromitsu (H); Tomita, Masahiro (M); Hino, Rika (R); Yoshizato, Katsutoshi (K); Mizuno, Shigeki (S); Tamura, Toshiki (T);

Affiliation: Insect Biotechnology and Sericology Department, National Institute of Agrobiological Sciences, 1-2 Owashi, Tsukuba, Ibaraki 305-8634, Japan.

Journal and publication information

Publication Type: Journal Article; Research Support, Non-U.S. Gov't

Journal: Insect biochemistry and molecular biology (Insect Biochem Mol Biol), published in England. (Language: eng)

Reference: 2005-Jan; vol 35 (issue 1) : pp 51-9

Dates: Created 2004/12/20; Completed 2005/04/21; Revised 2006/11/15;

PMID: 15607655, status: MEDLINE (last retrieval date: 2/18/2009, IMS Date: )

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MeSH headings (categories)

This article was linked to the MESH Headings shown below.

Animals
Bombyx - genetics; metabolism
Fibroins - biosynthesis; genetics
Gene Expression
Green Fluorescent Proteins - biosynthesis

Green Fluorescent Proteins - biosynthesis
Insect Proteins - biosynthesis; genetics
Mutation
Organisms, Genetically Modified
Recombinant Fusion Proteins - biosynthesis

Associated Chemicals: Insect Proteins (0) ; L-chain, fibroin protein, insect (0) ; Recombinant Fusion Proteins (0) ; Green Fluorescent Proteins (147336-22-9) ; Fibroins (9007-76-5)

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