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Competition Binding Between a Monoclonal Antibody and Serum Albumin for Several Lysophosphatidic Acid Isoforms in Solution
Author(s) -
Wojciak Jon,
Fleming Jonathan
Publication year - 2015
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.29.1_supplement.886.18
Subject(s) - lysophosphatidic acid , bovine serum albumin , gene isoform , receptor , monoclonal antibody , chemistry , biochemistry , human serum albumin , serum albumin , albumin , plasma protein binding , antibody , microbiology and biotechnology , biology , immunology , gene
Lysophosphatidic acid (LPA) is a bioactive signaling lipid that binds and activates specific G‐protein linked receptors on the cell surface to trigger numerous biological processes. LPA consists of multiple fatty acid isoforms with reportedly different receptor potencies. Serum albumin, an abundant carrier protein in mammals, is involved in the transport of LPA and may play a significant role in LPA‐receptor interactions. To further investigate this role, we used the solution based kinetic exclusion assay (KinExA) along with a monoclonal anti‐LPA antibody, B3, to measure the equilibrium dissociation constants (K D ) for bovine serum albumin (BSA) binding the most abundant native LPA species in human blood. Simultaneously, we were able to determine the K D for B3 binding these LPA species using “competition n‐curve analysis.” Our results show that BSA exhibits nanomolar affinities for these LPA species, and B3 has considerably higher affinity for each LPA isoform compared to BSA. We performed a subset of experiments using human serum albumin (HSA) and extrapolated that HSA has similar affinities for the same LPA species as BSA. These findings elucidate LPA‐protein interactions and describe a novel method to determine the K D for native lipids binding to proteins in solution.