Antibody crystallization on phospholipid films: dynamics and the effects of antibody conformation.
Monoclonal antibodies from two-dimensional (2-D) crystals when bound to haptenated phospholipid monolayers in physiological conditions and at ambient temperatures. IgG1 forms two crystal phases: a linear strand phase and a high-order hexagonal phase. The relative distribution of these two phases is dependent on temperature, pH, and salt concentration. This dependence is one that is associated with protein intramolecular interactions rather than lipid-lipid or lipid-protein interactions for a number of reasons: 1) Polyclonal antibodies against the hapten DNP do not organize into any crystal structure for any of the experimental conditions used. 2) Slightly denatured IgG (through storage at 4 degrees C, for example) does not readily crystallize and a shift in the temperature dependence for forming the hexagonal phase is observed. 3) There is no pH driven transition in crystallization tendency for IgE anti-DNP but a transition to disorder is observed at above 30 degrees C. No such transition exists for IgG1. Observation of the dynamics of crystal growth shows a clear and marked dependence on pH and temperature that is in accord with the results of long-term incubations. It is found that high pH retards crystal growth very significantly for IgG1 but not for IgE. Also, the crystal growth rate of 4 degrees C-stored IgG1 is greatly reduced over fresh IgG1 (-80 degrees C stored). Furthermore, it is found that the linear phase of IgG1 is an extremely rapidly forming phase but one that is metastable against the hexagonal phase.
D000818:Animals; D000911:Antibodies, Monoclonal; D003460:Crystallization; D004136:Dinitrobenzenes / Q000276:immunology; D004229:Dithiothreitol; D006863:Hydrogen-Ion Concentration; D007074:Immunoglobulin G; D051379:Mice; D008854:Microscopy, Electron / Q000379:methods; D010743:Phospholipids; D012965:Sodium Chloride; D013696:Temperature
D000911:Antibodies, Monoclonal; D004136:Dinitrobenzenes; D007074:Immunoglobulin G; D010743:Phospholipids; D012965:Sodium Chloride; D004229:Dithiothreitol