Testing the sulfotransferase molecular pore hypothesis

Ian Cook, Ting Wang, Steven C. Almo, Jungwook Kim, Charles N. Falany, Thomas S. Leyh

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Abstract

Human cytosolic sulfotransferases (SULTs) regulate the activities of hundreds of signaling metabolites via transfer of the sulfuryl moiety (-SO 3) from activated sulfate (3'-phosphoadenosine 5'-phosphosulfate) to the hydroxyls and primary amines of xeno- and endobiotics. How SULTs select substrates from the scores of competing ligands present in a cytosolic milieu is an important issue in the field. Selectivity appears to be sterically controlled by a molecular pore that opens and closes in response to nucleotide binding. This point of view is fostered by structures showing nucleotide-dependent pore closure and the fact that nucleotide binding induces an isomerization that restricts access to the acceptor-binding pocket. Molecular dynamics models underscore the importance of pore isomerization in selectivity and predict that specific molecular linkages stabilize the closed pore in response to nucleotide binding. To test the pore model, these linkages were disrupted in SULT2A1 via mutagenesis, and the effects on selectivity were determined. The mutations uncoupled nucleotide binding from selectivity and produced enzymes that no longer discriminated between large and small substrates. The mutations did not affect the affinity or turnover of small substrates but resulted in a 183-fold gain in catalytic efficiently toward large substrates. Models predict that an 11-residue "flap" covering the acceptor-binding pocket can open and admit large substrates when nucleotide is bound; a mutant structure demonstrated that this is so. In summary, the model was shown to be a robust, accurate predictor of SULT structure and selectivity whose general features will likely apply to other members of the SULT family.

Original languageEnglish (US)
Pages (from-to)8619-8626
Number of pages8
JournalJournal of Biological Chemistry
Volume288
Issue number12
DOIs
Publication statusPublished - Mar 22 2013

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ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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