Dissertation/Thesis Abstract

Human multidrug and toxin extrusion protein 1: Symmetry of substrate fluxes
by Dangprapai, Yodying, Ph.D., The University of Arizona, 2011, 140; 3460883
Abstract (Summary)

Human multidrug and toxin extrusion 1 (hMATE1) is a major candidate for being the molecular identity of organic cation/proton (OC/H+) exchange activity in the luminal membrane of renal proximal tubules (RPT). Although physiological function of hMATE1 supports luminal OC efflux, the kinetics of hMATE1-mediated OC transport have typically been characterized through measurement of uptake i.e., the interaction between outward-facing hMATE1 and OCs. To examine kinetics of hMATE1-mediated transport in a more physiologically relevant direction i.e., an interaction between inward-facing hMATE1 and cytoplasmic substrates, I measured the time course of hMATE1-mediated efflux of the prototypic MATE1-substrate, [3H]1-methyl-4-phenylpyridinium ([3H]MPP), under a variety of conditions, including different values for intra- and extracellular pH, from CHO cells that stably expressed hMATE1. I showed that an IC50/Ki for interaction between extracellular H+ and outward-facing hMATE1 determined from conventional uptake experiments [12.9 ± 1.23 nM (pH 7.89); n = 9] and from the efflux protocol [14.7 ± 3.45 nM (pH 7.83); n = 3] were not significantly different (P = 0.6). To test a hypothesis that H+ interacts symmetrically with each face of hMATE1, kinetics of interaction between intracellular H+ and inward-facing hMATE1 were determined using the efflux protocol. The IC50 for interaction with H+ was 11.5 nM (pH 7.91), consistent with symmetrical interactions of H+ with the inward-facing and outward-facing aspects of hMATE1. The efflux protocols demonstrated in this study are a potential means to examine kinetics at cytoplasmic face of hMATE1 and also a practical tool to screen uptake of substrates at extracellular face of hMATE1.

Indexing (document details)
Advisor: Wright, Stephen H.
Commitee: Brooks, Heddwen L., Dantzler, William H., Delamere, Nicholas A.
School: The University of Arizona
Department: Physiological Sciences
School Location: United States -- Arizona
Source: DAI-B 72/09, Dissertation Abstracts International
Subjects: Toxicology, Surgery, Pharmacology, Physiology
Keywords: Hmate1, Multidrug, Organic cation, Renal proximal tubule, Substrate fluxes, Toxin extrusion
Publication Number: 3460883
ISBN: 978-1-124-73567-2
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