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Expression and Characterization of a Variant of TACI (CRD2-shortTACIFc) in Pichia pastoris

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Abstract:

TACI is a member of the tumor necrosis factor receptor superfamily and serves as a key regulator of B cell function. The extracellular domain of a typical TNFR contains multiple copies of CRD, which bind in the monomermonomer interfaces of a trimeric ligand. TACI binds to two ligands, APRIL and BAFF, with high affinity and contains two CRD in its extracellular regions, while BCMA and BR3, contain a single or partial CRD for binding the two ligands. However, TACI can be classified as a single CRD receptor because the amino-terminal CRD1 doesn't contribute to ligand binding. To obtain a new variant of TACI possessing higher affinities for binding, we fused a repeat sequence of CRD2 to the N-terminus of the short form of TACI. The new APRIL antagonist peptide, CRD2-shortTACI-Fc, was designed based on the modeling 3-D complex structure of TACI and APRIL. As expected, the purified recombinant CRD2-shortTACI-Fc fusion protein could bind to APRIL in vitro and demonstrated dose-dependent inhibition of APRIL-induced proliferative activity in Raji cells. We found that CRD2-shortTACI-Fc, has a higher affinity for binding to ligands than short-TACI-Fc, which contains a single CRD2.





Keywords: APRIL; BAFF; CRD2; DNA ligase; Novagen; Pichia pastoris; TACI; affinity; autoimmune disorders; homology modeling

Document Type: Research Article

DOI: http://dx.doi.org/10.2174/092986612799363136

Publication date: March 1, 2012

More about this publication?
  • Protein & Peptide Letters publishes short papers in all important aspects of protein and peptide research, including structural studies, recombinant expression, function, synthesis, enzymology, immunology, molecular modeling, drug design etc. Manuscripts must have a significant element of novelty, timeliness and urgency that merit rapid publication. Reports of crystallisation, and preliminary structure determinations of biologically important proteins are acceptable. Purely theoretical papers are also acceptable provided they provide new insight into the principles of protein/peptide structure and function.
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