Mechanisms of lymphocyte activation. Binding kinetics of phytohemagglutinin to human lymphocytes.
The interactions of phytohemagglutinin (PHA) with normal human lymphocytes were studied utilizing radioiodinated leukoagglutinin (125I-LPHA) over a concentration spectrum encompassing the entire range of lymphocyte metabolic responses. 125I-LPHA binding was temperature-, pH-, and time-dependent. Ligand association was rapid with a t1/2 of 3 to 5 min, reaching steady state in 30 min at 22 degrees. Receptor specificity was demonstrated by the high receptor affinity for 125I-LPHA and by quantitative inhibition of 125I-LPHA binding with LPHA and 127I-LPHA but not with concanavalin A or bovine serum albumin. Under our experimental conditions there was no measurable degradation of 125I-LPHA and no detectable shedding of 125I-LPHA receptors or receptor-125I-LPHA complexes. Equilibrium studies of 125I-LPHA interactions with specific lymphocyte membrane receptors generated a complex curvilinear Scatchard plot. This, added to progressive deceleration of the dissociation reaction inversely proportional to receptor occupancy by 125I-LPHA, reflects changing receptor affinity for the ligand and suggests site-site interactions of the negative cooperativity type. These interactions which appear to be common to all lymphocyte subpopulations, preclude accurate calculation of lymphocyte binding capacity for 125I-LPHA and of physically meaningful affinity constants. Although the fate and role of a small fraction of apparently nondissociable 125I-LPHA remains to be elucidated, occupancy-dependent receptor affinity for 125I-LPHA, dissociation of receptor-125I-LPHA complexes, retention of binding properties by cell-exposed 125I-LPHA, and the large numbers of spare surface receptors for 125I-LPHA might represent important mechanisms for modulating cell activation by 125I-LPHA.
D001665:Binding Sites; D006801:Humans; D007700:Kinetics; D037102:Lectins; D008213:Lymphocyte Activation; D008214:Lymphocytes / Q000502:physiology*; D013696:Temperature
D037102:Lectins