Differences in the substrate binding regions of renal organic anion transporters 1 (OAT1) and 3 (OAT3)

Bethzaida Astorga, Theresa M. Wunz, Mark Morales, Stephen H. Wright, Ryan M. Pelis

Research output: Contribution to journalArticle

10 Scopus citations

Abstract

This study examined the selectivity of organic anion transporters OAT1 and OAT3 for structural congeners of the heavy metal chelator 2,3-dimercapto-1-propanesulfonic acid (DMPS). Thiol-reactive reagents were also used to test structural predictions based on a homology model of OAT1 structure. DMPS was near equipotent in its ability to inhibit OAT1 (IC50 = 83 (μM) and OAT3 (IC50 = 40 (μM) expressed in Chinese hamster ovary cells. However, removal of a thiol group (3-mercapto-1-propanesulfonic acid) resulted in a 2.5-fold increase in IC50 toward OAT1 vs. a ~55-fold increase in IC50 toward OAT3. The data suggested that compound volume/size is important for binding to OAT1/OAT3. The sensitivity to HgCl2 of OAT1 and OAT3 was also dramatically different, with IC50 values of 104 and 659 (μM, respectively. Consistent with cysteines of OAT1 being more accessible from the external medium than those of OAT3, thiol-reactive reagents reacted preferentially with OAT1 in cell surface biotinylation assays. OAT1 was less sensitive to HgCl2 inhibition and less reactive toward membrane-impermeant thiol reactive reagents following mutation of cysteine 440 (C440) to an alanine. These data indicate that C440 in transmembrane helix 10 of OAT1 is accessible from the extracellular space. Indeed, C440 was exposed to the aqueous phase of the presumptive substrate translocation pathway in a homology model of OAT1 structure. The limited thiol reactivity in OAT3 suggests that the homologous cysteine residue (C428) is less accessible. Consistent with their homolog-specific selectivities, these data highlight structural differences in the substrate binding regions of OAT1 and OAT3.

Original languageEnglish (US)
Pages (from-to)F378-F386
JournalAmerican Journal of Physiology - Renal Physiology
Volume301
Issue number2
DOIs
StatePublished - Aug 1 2011

Keywords

  • Cysteine accessibility
  • Heavy metal chelator
  • Homology modeling
  • Proximal tubule

ASJC Scopus subject areas

  • Physiology
  • Urology

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