Expression in insect cells of the functional domain of CD21 (complement receptor type two) as a truncated soluble molecule using a baculovirus vector

Prodinger,W.M.; Schoch,J.; Schwendinger,M.G.; Hellwage,J.; Parson,W.; Zipfel,P.F.; Dierich,M.P.
We have made use of the plasmid vector pBSV-8His recently established for baculovirus-mediated expression of His-tagged proteins for the production of a truncated soluble complement regulator protein. The protein comprised the N-terminal part, i.e. short consensus repeats (SCRs) 1-4, of the B-cell membrane protein complement receptor type two (CR2; CD21) and contained the functional epitopes which mediate the binding of the complement component C3 fragments C3dg and iC3b. This recombinant protein, termed rsCR2.1-4, was furnished with a C-terminal histidine-tag for easy purification from insect cell supernatant. The yield of > 90% pure rsCR2.1-4 was 3 micrograms/ml supernatant at day eight p.i. RsCR2.1-4 was expressed as two proteins with a M(r) of 29 and 31 representing differentially glycosylated forms. Both reacted specifically with anti-CR2 mAb HB5 directed against SCRs 3-4, but not with anti-CR2 mAbs recognizing SCRs beyond SCR 4. RsCR2.1-4 was able to bind C3dg and to block binding of C3dg-coated beads to Raji cells. Used as antigen for immunization, it allowed the efficient and well-aimed generation of antisera which specifically blocked attachment of C3dg-coated beads to Raji B cells. Thus, insect cell derived rsCR2.1-4 has proved a valuable tool to study the functional domain of CR2 and its immunoregulatory capacity in B-lymphocytes
Immunopharmacology 1997 38(1-2):141-148
Tags: Antibodies,Monoclonal; Antigen-Antibody Reactions; Austria; B-Lymphocytes; Baculoviridae; biosynthesis; female; Genetic Vectors; genetics; human; Immunization; immunology; metabolism; Mice; Mice,Inbred BALB C; Plasmids; Receptors,Complement; Receptors,Complement 3d; Recombinant Proteins; Spodoptera
PubMed: 9476125
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