Journal Article

Characterization of SpsQ from Staphylococcus pseudintermedius as an affinity chromatography ligand for canine therapeutic antibodies

Takeuchi, Hiroto, Murata, Shiro, Ohashi, Kazuhiko, Suzuki, Yasuhiko, Maekawa, Naoya, Okagawa, Tomohiro, Usui, Masaru, Nakajima, Chie, Konnai, Satoru, Tamura, Yutaka

2023

Overview

Coagulase-positive Staphylococci express protein A, which binds to host antibodies, to evade the immune system. Taking advantage of its specific binding to antibodies, protein A from Staphylococcus aureus , which is called SpA, is commonly used as an affinity chromatography ligand for human therapeutic antibodies. However, among four canine IgG subclasses (A, B, C, and D), only IgG-B binds to SpA strongly and establishing an efficient and robust purification scheme for canine therapeutic antibodies whose IgG subclass is A, C, or D remains difficult and depends on finding a suitable substitute to SpA. S . pseudintermedius , a major coagulase-positive Staphylococci found in dogs, expresses spsQ gene which is orthologous to S . aureus spa . We hypothesized that to serve S . pseudintermedius to better adapt to the dog immune system, SpsQ would bind to canine IgGs stronger than SpA, making it a better affinity chromatography ligand for canine therapeutic antibodies. To characterize SpsQ, we first determined the spsQ nucleotide sequence from S . pseudintermedius isolates. Based on the identified sequence, we prepared recombinant proteins containing the immunoglobulin-binding domains of SpA (r-SpA) and SpsQ (r-SpsQ) and determined their binding capacity for each canine IgG subclass. The binding capacity of r-SpsQ for IgG-B was almost as high as that of r-SpA. Interestingly, while both r-SpsQ and r-SpA showed no binding to IgG-C, the binding capacity of r-SpsQ for IgG-A and IgG-D was significantly higher than that of r-SpA. Finally, we performed affinity chromatography using r-SpsQ- or r-SpA-immobilized resin and revealed that the recovery rates of IgG-A and IgG-D using r-SpsQ were significantly higher than those using r-SpA. Our findings indicate that SpsQ has a strong potential to be used as an affinity chromatography ligand for canine therapeutic antibodies of subclass A, B, and D.

• Publisher: Public Library of Science,Public Library of Science (PLoS)
• Subject: Affinity / Affinity chromatography / Animals / Antibodies / Bacterial proteins / Binding / Biology and Life Sciences / Biopharmaceutics / Carbenicillin / Chromatography / Chromatography, Affinity / Cloning / Coagulase / Cytotoxicity / Diseases / Dogs / Ethylenediaminetetraacetic acid / Genetic aspects / Health aspects / Immune system / Immunoglobulin G / Immunologic Factors / Ligands / Ligands (Biochemistry) / Medicine and Health Sciences / Nucleotide sequence / Nucleotides / Pharmaceutical industry / Physiological aspects / Protein A / Protein binding / Proteins / Recombinant proteins / Research and Analysis Methods / Scientific equipment and supplies industry / Staphylococcal Protein A - metabolism / Staphylococcus / Staphylococcus aureus / Staphylococcus aureus - metabolism / Staphylococcus pseudintermedius / Structure / Viral antibodies