Active Site Gating Controls Substrate Selectivity in Cytosolic Sulfotransferases

Abstract

Human cytosolic sulfotransferases (SULTs) transfer the sulfuryl‐moiety (‐SO3) from activated sulfate (PAPS, 3′‐phosphoadenosine 5‐phosphosulfate) to the hydroxyls and primary amines of metabolites and xenobiotics. The addition of the sulfuryl‐group radically alters the acceptor‐target interactions. SULT1A1 and 2A1, which comprise ~90% of the liver SULTs by mass, serve two critical functions in the liver. First is regulating signaling hormones to maintain homeostasis. The homeostasis function requires a high affinity to a set of conserved substrates. The second function is protecting the body from xenobiotics that can disrupt receptor signaling. The protective function requires a broader substrate specificity to handle the diversity of compounds in the environment. In this work we expand on our proposed mechanism for substrate selectivity where the binding of PAPS induces an isomerization that opens and closes the “cap” of the active site. This mechanism allows the SULT to sulfate a wide range of substrates, while maintaining homeostatic function. Computer modeling identified three interactions that regulate the gating. These interactions were tested using mutagenesis. The mutants were examined using equilibrium and pre‐steady state binding assays to measure the influence to gating. All of the mutants resulted in the loss of gating and substrate selectivity, confirming the model.