Monomeric and dimeric chemokine-Id fusion proteins and their genetic construction. (Left) Schematic structure of monomeric chemokine-Id fusion protein and the corresponding gene construct. The mouse MIP-1α chemokine gene, or a control construct that encodes MIP-1α with point mutations replacing the first cysteine with serine (C11S), was fused in frame with the scFv, separated by a spacer fragment (NDAQAPKS) to enable proper folding of the proteins.¹³  (Right) Structure of homodimeric chemokine-Id fusion protein (vaccibody) and the corresponding gene constructs. Targeting, dimerization, and antigenic units are indicated, as are variants in the different units (see also Table S1). In the monomeric chemokine-scF_v constructs, the MIP-1α and RANTES mouse chemokine genes, or variants where the first cysteine had been destroyed, were inserted in the targeting cassette of the pLNOH2 vector. In the vaccine constructs with a dimerization unit derived from mouse IgG2b, the single hinge exon (4 cysteines that can form disulphide bonds) was fused in frame with a G₃S₂G₃SG linker (linker 1) and the C_γ3 exon. A short GLGGL linker (linker 2) connects the C_γ3 exon with the antigenic scFv from either MOPC315 myeloma or the A20 B-cell lymphoma. In the constructs with a dimerization unit derived from human IgG3, there is an intronic sequence upstream of the h1 hinge exon, which is fused in frame with the h4 exon (altogether 3 cysteines that can form disulphide bonds) and linker 1 followed by the C_γ3 exon, a GLGGL sequence (linker 2) and scFv. A (G₄S)₃ linker (linker 3) connects the V_H and V_L in the antigenic unit.