Kinetic analysis of ligand binding to the Ehrlich cell nucleoside transporter: Pharmacological characterization of allosteric interactions with the sup 3 Hnitrobenzylthioinosine binding site

Journal Article · Sat Jun 01 04:00:00 EDT 1991 · Molecular Pharmacology; (United States)

OSTI ID:5556931

Hammond, J R ^[1]

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1. Department of Pharmacology and Toxicology, University of Western Ontario, London (Canada)

Kinetic analysis of the binding of {sup 3}Hnitrobenzylthioinosine ({sup 3}H NBMPR) to Ehrlich ascites tumor cell plasma membranes was conducted in the presence and absence of a variety of nucleoside transport inhibitors and substrates. The association of {sup 3}H NBMPR with Ehrlich cell membranes occurred in two distinct phases, possibly reflecting functional conformation changes in the {sup 3}HNBMPR binding site/nucleoside transporter complex. Inhibitors of the equilibrium binding of {sup 3}HNBMPR, tested at submaximal inhibitory concentrations, generally decreased the rate of association of {sup 3}HNBMPR, but the magnitude of this effect varied significantly with the agent tested. Adenosine and diazepam had relatively minor effects on the association rate, whereas dipyridamole and mioflazine slowed the rate dramatically. Inhibitors of nucleoside transport also decreased the rate of dissociation of {sup 3}HNBMPR, with an order of potency significantly different from their relative potencies as inhibitors of the equilibrium binding of {sup 3}HNBMPR. Dilazep, dipyridamole, and mioflazine were effective inhibitors of both {sup 3}HNBMPR dissociation and equilibrium binding. The lidoflazine analogue R75231, on the other hand, had no effect on the rate of dissociation of {sup 3}HNBMPR at concentrations below 300 microM, even though it was one of the most potent inhibitors of {sup 3}HNBMPR binding tested (Ki less than 100 nM). In contrast, a series of natural substrates for the nucleoside transport system enhanced the rate of dissociation of {sup 3}HNBMPR with an order of effectiveness that paralleled their relative affinities for the permeant site of the transporter. The most effective enhancers of {sup 3}HNBMPR dissociation, however, were the benzodiazepines diazepam, chlordiazepoxide, and triazolam.

• OSTI ID: 5556931
• Journal Information: Molecular Pharmacology; (United States), Journal Name: Molecular Pharmacology; (United States) Vol. 39:6; ISSN MOPMA; ISSN 0026-895X
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AFFINITY
ANIMAL CELLS
ANIMALS
AROMATICS
AZAARENES
BIOCHEMICAL REACTION KINETICS
CELL CONSTITUENTS
CELL MEMBRANES
CENTRAL NERVOUS SYSTEM AGENTS
CHEMICAL COMPOSITION
DRUGS
HETEROCYCLIC COMPOUNDS
HYDROGEN COMPOUNDS
INHIBITION
INOSINE
ISOTOPE APPLICATIONS
KINETICS
LIGANDS
MAMMALS
MEMBRANE PROTEINS
MEMBRANE TRANSPORT
MEMBRANES
MICE
NUCLEOSIDES
NUCLEOTIDES
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
PHARMACOLOGY
PROTEINS
PSYCHOTROPIC DRUGS
PURINES
REACTION KINETICS
RECEPTORS
RIBOSIDES
RODENTS
TRACER TECHNIQUES
TRANQUILIZERS
TRITIUM COMPOUNDS
TUMOR CELLS
VERTEBRATES