title: Temperature differentially affects encounter and docking thermodynamics of antibody--antigen association.
abstract: Using BIACORE SPR, we have examined the mechanism of temperature effects on the binding kinetics of two closely related antibody Fabs (H10 and H26) which recognize coincident epitopes on hen egg-white lysozyme (HEL), and whose association and dissociation kinetics are best described by the two-step conformational change model which we interpret as molecular encounter and docking. Time-course series data obtained at a series of six temperatures (6, 10, 15, 25, 30 and 37 degrees C) showed that temperature differentially affects the rate constants of the encounter and docking steps. Docking is more temperature-sensitive than the encounter step, and energetically less favorable at higher temperatures. At elevated temperatures, the time required for docking is longer and the apparent increase in off-rate reflects the greater proportion of the molecules failing to dock and remaining in the less stable encounter state. As a consequence, distribution of free energy change between the encounter and docking steps is altered. At physiological temperature (37 degrees C) the docking step of the H26 complex is energetically unfavorable and most complexes essentially do not dock. There is a significant decrease in total free energy change of the H26 complex at higher temperatures. Elevated temperature changes the rate-limiting step of H26--HEL association from the encounter to the docking step, but not that of H10--HEL. Our results indicate that the mechanism by which elevated temperature reduces the affinities of antigen--antibody complexes is to decrease the net docking rate, and/or stability of the docked complex; at higher temperatures, a smaller proportion of the complexes actually anneal to a more stable docked state. This mechanism may have broad applicability to other receptor--ligand complexes.
authors.affiliation: Basic Research Laboratory, National Cancer Institute, Frederick, MD 21702-1201, USA.
authors.forename: Claudia A
authors.initials: CA
authors.lastname: Lipschultz
authors.forename: Anthony
authors.initials: A
authors.lastname: Yee
authors.forename: S
authors.initials: S
authors.lastname: Mohan
authors.forename: Yili
authors.initials: Y
authors.lastname: Li
authors.forename: Sandra J
authors.initials: SJ
authors.lastname: Smith-Gill
mesh_terms: D000936:Antigen-Antibody Complex; D000939:Epitopes / Q000737:chemistry / Q000276:immunology; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry* / Q000276:immunology; D009113:Muramidase / Q000737:chemistry* / Q000276:immunology; D013696:Temperature; D013816:Thermodynamics
chemical_list: D000936:Antigen-Antibody Complex; D000939:Epitopes; D007140:Immunoglobulin Fab Fragments; C421864:hen egg lysozyme; D009113:Muramidase