[Cloning, soluble expression and characterization of human sBCMA]

Area of Science:

• Biochemistry
• Molecular Biology
• Immunology

Background:

• B-cell maturation antigen (BCMA) is a transmembrane receptor involved in B-cell regulation.
• Soluble BCMA (sBCMA) is being investigated as a potential decoy receptor to modulate B-cell responses.
• Structural and functional characterization requires active, recombinant sBCMA.

Purpose of the Study:

• To establish a feasible method for expressing active soluble BCMA (sBCMA).
• To obtain recombinant sBCMA for structural and functional research.
• To validate the bioactivity of the expressed recombinant sBCMA.

Main Methods:

• Full-length hBCMA was amplified via RT-PCR from Raji cell line RNA.
• The cDNA for the extracellular soluble domain of hBCMA was cloned into a pET43.1a(+) vector.
• The recombinant vector was transformed into E. coli Origami B(DE3) pLyS for disulfide bond formation.
• Recombinant protein (sBCMA-NusA-His6) was expressed, identified by Western blotting, and purified using Ni(+)-chelating Sepharose.
• Binding activity with BAFF was assessed by ELISA, and B-cell proliferation inhibition was evaluated.

Main Results:

• A soluble fusion protein, sBCMA-NusA-His6, was successfully expressed and purified.
• Recombinant sBCMA demonstrated binding activity with BAFF.
• Recombinant sBCMA effectively inhibited the proliferation of mouse B cells stimulated by rhsBAFF.
• The expression method proved feasible and effectual for proteins rich in disulfide bonds.

Conclusions:

• The developed method enables the production of bioactive recombinant soluble BCMA (sBCMA).
• This recombinant sBCMA is suitable for further structural and functional investigations.
• The expression strategy is effective for producing disulfide-rich proteins.