Inhibition of the human sodium/bile acid cotransporters by side-specific methanethiosulfonate sulfhydryl reagents: Substrate-controlled accessibilityof site of inactivation

Mammalian sodium/bile acid cotransporters (SBATs) constitute a subgroup of the sodium cotransporter superfamily and function in the enterohepatic circ ulation of bile acids. They are glycoproteins with an exoplasmic N-terminus , seven or nine transmembrane segments, and a cytoplasmic C-terminus. They exhibit no significant homology with other members of the sodium cotranspor ter family and there is limited structure/function information available fo r the SBATs. Membrane-impermeant methanethiosulfonates (MTS) inhibited bile acid transport by alkylation of cysteine 270 (apical SBAT)/266 (basolatera l SEAT) that is fully conserved among the sodium/bile acid cotransporters. The accessibility of this residue to MTS reagent is regulated by the natura l substrates, sodium and bile acid. In experiments with the apical SEAT, so dium alone increases the reactivity with the thiol reagents as compared to sodium-free medium. In contrast, bile acids protect the SBATs from inactiva tion, although only in the presence of sodium, The inhibition and protectio n data suggest that cysteine 270/266 lies in a sodium-sensitive region of t he SBATs that is implicated in bile acid transport.