Synthesis and characterization of chimeric silkworm silk.
A synthetic gene encoding a chimeric silklike protein was constructed that combined a polyalanine encoding region (Ala)(18), a sequence slightly longer than the (Ala)(12-13) found in the silk fibroin from the wild silkworm Samia cynthia ricini, and a sequence encoding GVGAGYGAGAGYGVGAGYGAGVGYGAGAGY, found in the silk fibroin from the silkworm Bombyx mori. A tetramer of the chimeric repeat sequence encoding a approximately 29 kDa protein was expressed as a fusion protein in Escherichia coli. In comparison to S. c. ricini silk, the chimeric protein demonstrated improved solubility because it could be dissolved in 8 M urea. The purified protein assumed an alpha-helical structure based on solid-state (13)C CP/MAS NMR and was less prone to conformational transition to a beta-sheet, unlike native silk proteins from S. c. ricini. Model peptides representing the crystalline region of S. c. ricini silk fibroin, (Ala)(12) and (Ala)(18), formed beta-sheet structures. Therefore, the solubility and structural transitions of the chimeric protein were significantly altered through the formation of this chimeric silk. This experimental strategy to the study of silk structure and function can be used to develop an improved understanding of the contributions of protein domains in repetitive silkworm and spider silk sequences to structure development and structural transitions.
Asakura
Tetsuo
T
Department of Biotechnology, Tokyo University of Agriculture and Technology, Koganei, Tokyo 184-8588, Japan. asakura@cc.tuat.ac.jp
Nitta
Koji
K
Yang
Mingying
M
Yao
Juming
J
Nakazawa
Yasumoto
Y
Kaplan
David L
DL
D000595:Amino Acid Sequence / Q000502:physiology; D000818:Animals; D001483:Base Sequence / Q000502:physiology; D012831:Bombyx / Q000235:genetics / Q000378:metabolism*; D002417:Cattle; D019476:Insect Proteins / Q000096:biosynthesis* / Q000235:genetics; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics; D047011:Silk
D019476:Insect Proteins; D011993:Recombinant Fusion Proteins; D047011:Silk