Human pIgR mimetic peptidic ligand for affinity purification of IgM: Part I: Ligand design and binding mechanism

Abstract

Synthetic peptides incorporating the complementarity determining region 2 (CDR2)-like loop of domain 1 of human polymeric immunoglobulin receptor (hpIgR-D1) were investigated for their interactions with human Immunoglobulin-M (hIgM) and other hIgs. pep14, a 14mer peptide, emerged as a unique biomimetic ligand with specificity to interact with hIgM and negligible affinity for hIgG, hIgE, hIgA1 and bovine serum albumin. Surface plasmon resonance-based binding studies between immobilized pep14 and fragments of hIgM molecule, namely Fc5μ and Fab, suggested that pep14 was binding to a motif in the constant domain 2 of hIgM. Modified peptides, pep13, pep5A and pep14A were investigated for their interaction with hIgM/ hIgG1 to elucidate the binding mechanism of pep14. Cysteine in pep14 was identified to be crucial for inducing thiophilic-like interactions between the ligand and hIgM while glutamic acid had a significant role in attributing specificity to the ligand to interact with hIgM. Circular dichroism studies suggested the absence of native β-hairpin structure in pep14. The ligand exhibited a more flexible conformation consisting mainly of a mixture of coil and turn.