Pressure-Induced Dissociation of Antigen-Antibody Complexes

Abstract

Pressures on the order of 1000-4000 bar have been reported to reversibly dissociate a number of oligomeric protein complexes without gross changes in protein structure. Here, we report that hydrostatic pressure can also dissociate some antigen-antibody complexes in solution. The association of fluorescent-labeled antigens with monoclonal antibodies was monitored via increases in the fluorescence anisotropy upon binding. Previously, we had found that pressures of 2000 atm were able to dissociate bovine serum albumin (BSA) from immunoadsorbents formed from certain antibodies but not others. In this study, we have found that the sensitivity to pressure in solution is present for the interaction of BSA with MAb 9.1 and absent for the interaction of BSA with MAb 6.1; this behavior is consistent with the immunoadsorbent study. The interaction of hen egg white lysozyme with two monoclonal antibodies was also measured. Interestingly, the complex with the greater electrostatic character (HyHEL-5) did not exhibit pressure sensitivity, as would be expected due to electrostriction effects, whereas the more hydrophobic complex (HyHEL-10) exhibited a strong pressure sensitivity. In each of the systems displaying pressure sensitivity, the free energy of association was found to increase linearly with pressure, indicating a constant change in volume between the free and bound states. Overall, these results indicate that some antigen-antibody complexes exhibit significant sensitivity to pressure, whereas others do not; the mechanisms that discriminate between these cases remain unresolved. Understanding and manipulation of this phenomenon may prove useful in a variety of processes involving the recovery from antigens of antibodies.

Inhibition of anti-fluorescent probe monoclonal antibody by long-chain amphiphiles

“…The quenching decreased with pressure as seen in the figure. In the previous paper [24], we have calculated the standard volume change associated with the dissociation of FL and IgG-49 from the FL-IgG-49 complex and obtained the value of −5.2 cm 3 mol −1 , that is, the complex expands slightly by binding of them. Similar dissociation behavior of anti-fluorescyl [25][26][27] and anti-BSA [28] monoclonal antibodies by pressure has been observed, and our value is close to the value reported for the anti-fluorescyl monoclonal antibody with a strong affinity to FL by Herron et al [15]. The value decreased with increasing temperature.…”

Single‐domain antibody fragments with high conformational stability

“…Therefore, in this pressure range, it should be feasible to disrupt antigen–V H H complexes without unfolding either molecule. This strategy can be advantageously used for immunoaffinity separation, where a specifically bound antigen is eluted from the immunoadsorbent after a controlled pressure increase (Olson et al 1989; Sudaram et al 1998). Compared with the harsh conditions usually used, such a “hyperbaric elution” extends the immunoadsorbent lifetime (Olson et al 1989).…”

High Hydrostatic Pressure Can Probe the Effects of Functionally Related Ligands on the Quaternary Structures of the Chaperonins GroEL and GroES

“…The rates were evaluated by fitting the data to either mono-or biexponential equations: 3 , respectively. The independent variable Y was the observed scattering intensity in counts/s after subtracting the scattering due to buffer.…”

“…Their quaternary structures are crucial to the mechanism of chaperoninassisted protein folding. These two proteins are coexpressed from a common GroE operon in Escherichia coli (1)(2)(3). Mutational studies have demonstrated that both chaperonins are essential for protein folding in vivo (4 -6).…”