Pressure-induced conformational changes in an antigen and an antibody and the implications on their use for hyperbaric immunoadsorption.
Pressure-induced conformational changes in two proteins, bovine serum albumin (BSA) and immunoglobulin G (IgG), were studied to assess the application of hyperbaric manipulation to the dissociation of antigen-antibody complexes. Antigen-antibody dissociation is important in the product-recovery phase of immunoadsorption, an affinity purification process. Three techniques were used in parallel for this study, including fluorescence, Fourier transform infrared (FTIR) spectroscopy, and the enzyme-linked immunosorbent assay (ELISA). Employing a fluorescent probe, fluorescent intensity measurements were used to detect protein conformational changes. FTIR spectroscopy was used to determine changes in protein secondary structure induced by high pressure, while the ELISA test was used to examine antibody recognition after the proteins had been pressure-treated. The results from this work demonstrate that IgG is resistant to conformational changes induced by pressures below 2 kbar. In contrast, BSA undergoes reversible conformational changes in this pressure range. However, these conformational changes are not reflected in tests measuring antibody recognition. These findings indicate that IgGs have the potential to be used as recycled ligands in immunoadsorption separation processes. Different antigens that are being considered for purification by immunoadsorption and separated by means of high pressure could be screened by the methods disclosed to determine their stability under high pressure conditions.
Howlett
J R
JR
Department of Biology, Carleton University, Ottawa, Ontario, Canada.
Ismail
A A
AA
Armstrong
D W
DW
Wong
P T
PT
D000327:Adsorption; D000936:Antigen-Antibody Complex; D004797:Enzyme-Linked Immunosorbent Assay; D005453:Fluorescence; D006874:Hydrostatic Pressure; D007074:Immunoglobulin G / Q000737:chemistry*; D011487:Protein Conformation; D012710:Serum Albumin, Bovine / Q000737:chemistry*; D013055:Spectrophotometry, Infrared
D000936:Antigen-Antibody Complex; D007074:Immunoglobulin G; D012710:Serum Albumin, Bovine