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Uptake and destruction of 125 I‐CSF‐1 by peritoneal exudate macrophages
Author(s) -
Guilbert Larry J.,
Tynan P. Wendy,
Stanley E. Richard
Publication year - 1986
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/jcb.240310303
Subject(s) - internalization , dissociation (chemistry) , ligand (biochemistry) , reaction rate constant , chemistry , dissociation constant , biophysics , kinetics , stereochemistry , biochemistry , cell , receptor , biology , physics , quantum mechanics
The binding and uptake of the colony‐stimulating factor CSF‐1 by peritoneal exudate macrophages (PEM) from lipopolysaccharide insensitive C3H/HeJ mice was examined at 2°C, and at 37°C. At 2°C, 125 I‐CSF‐1 was bound irreversibly to the cell surface. At 37°C, 90% of the cell surface associated 125 I‐CSF‐1 was rapidly internalized and subsequently degraded and the remaining 10% dissociated as intact 125 I‐CSF‐1. Thus classical equilibrium or steady state methods could not be used to quantitatively analyze ligand–cell interactions at either temperature, and alternative approaches were developed. At 2°C, the equilibrium constant (K d ⩽ 10 −13 M) was derived from estimates of the rate constants for the binding (k on ≃ 8 × 10 5 M −1 s −1 ) and dissociation (k off ⩽ 2 × 10 −7 s −1 ) reactions. At 37°C, the processes of dissociation and internalization of bound ligand were kinetically competitive, and the data was formally treated as a system of competing first order reactions, yielding first order rate constants for dissociation, k off = 0.7 min −1 (t 1/2 = 10 min) and internalization, k in = 0.07 min −1 (t 1/2 = 1 min). Approximately 15 min after internalization, low‐molecular weight 125 I‐labeled degradation products began to appear in the medium. Release of this degraded 125 I‐CSF‐1 was kinetically first order over three half‐lives (K d = 4.3 × 10 −2 min −1 , t 1/2 = 16 min). Thus CSF‐1 binds to a single class of receptors on PEM, is internalized with a single rate limiting step, and is rapidly destroyed without segregation into more slowly degrading intracellular compartments.