Isolation, characterization, and cDNA sequence of a carotenoid binding protein from the silk gland of Bombyx mori larvae.
A carotenoid binding protein (CBP) has been isolated from the silk glands of Bombyx mori larvae. The protein has an apparent molecular mass of 33 kDa and binds carotenoids in a 1:1 molar ratio. Lutein accounts for 90% of the bound carotenoids, whereas alpha-carotene and beta-carotene are minor components. Immunological analysis demonstrated the presence of CBP only in the yellow-colored tissues of the silk gland, midgut, testis, and ovary. Several phenotypes of B. mori mutants linked to carotenoid transport have been utilized to characterize CBP. The Y (yellow hemolymph) gene controls uptake of carotenoids from the midgut lumen into the midgut epithelium, and larvae with the +(Y) gene lack this property. Immunoblotting analysis confirmed the presence of CBP in mutants with the dominant Y gene only. Immunohistochemistry verified the localization of CBP in the villi of the midgut epithelium, indicating that CBP might be involved in absorption of carotenoids. A cDNA clone for CBP encoding a protein of 297 amino acids has been isolated from the B. mori silk gland cDNA library. The deduced amino acid sequence revealed that CBP is a novel member of the steroidogenic acute regulatory (StAR) protein family with its unique structural feature of a StAR-related lipid transfer domain, known to aid in lipid transfer and recognition. Lutein-binding capacity of the recombinant CBP (rCBP) determined by incubating rCBP with lutein followed by immunoprecipitation using anti-CBP IgG conjugated to protein A-Sepharose, demonstrated the formation of a lutein-rCBP complex. Sequence analyses coupled with binding specificity suggest that CBP is a new member of the StAR protein family that binds carotenoids rather than cholesterol.
Tabunoki
Hiroko
H
Bio-Application and System Engineering, Tokyo University of Agriculture and Technology, Koganei, Tokyo 183-0054, Japan.
Sugiyama
Hiromu
H
Tanaka
Yoshiro
Y
Fujii
Hiroshi
H
Banno
Yutaka
Y
Jouni
Zeina E
ZE
Kobayashi
Masahiko
M
Sato
Ryoichi
R
Maekawa
Hideaki
H
Tsuchida
Kozo
K
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D012831:Bombyx / Q000235:genetics* / Q000502:physiology; D002352:Carrier Proteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D005088:Exocrine Glands / Q000378:metabolism*; D019476:Insect Proteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D007814:Larva; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016296:Mutagenesis; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D002352:Carrier Proteins; D018076:DNA, Complementary; D019476:Insect Proteins