Paradigms of Sulfotransferase Catalysis

Wang, Ting; Cook, Ian; Falany, Charles N.; Leyh, Thomas S.

doi:10.1074/jbc.m114.573501

Cited by 39 publications

(77 citation statements)

References 52 publications

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“…Sulfonation has been shown to be a high-affinity, low-capacity conjugation reaction (70,71) and PAPS availability might influence the level of catalytic activity (72). Also, PAPS release was determined to be a rate-limiting step during sulfonation reactions (72) and "trapped," un-sulfonated cofactor (PAP) could promote the formation of catalytically incompetent dead-end complexes (PAP-enzyme-ligand). These fluctuations in cytosol composition, molecule concentrations, and also the dynamics of cofactor release remain challenging to reproduce in a computerbased prediction model.…”

Section: Discussionmentioning

confidence: 99%

“…In vivo concentrations of PAPS can significantly vary depending on the tissue and depending on sulfonation rates of other heavily sulfonated molecules present in the cytosol (55). Sulfonation has been shown to be a high-affinity, low-capacity conjugation reaction (70,71) and PAPS availability might influence the level of catalytic activity (72). Also, PAPS release was determined to be a rate-limiting step during sulfonation reactions (72) and "trapped," un-sulfonated cofactor (PAP) could promote the formation of catalytically incompetent dead-end complexes (PAP-enzyme-ligand).…”

Section: Discussionmentioning

confidence: 99%

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In Silico Prediction of Human Sulfotransferase 1E1 Activity Guided by Pharmacophores from Molecular Dynamics Simulations

Rakers

Schumacher

Meinl

et al. 2016

Journal of Biological Chemistry

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Acting during phase II metabolism, sulfotransferases (SULTs) serve detoxification by transforming a broad spectrum of compounds from pharmaceutical, nutritional, or environmental sources into more easily excretable metabolites. However, SULT activity has also been shown to promote formation of reactive metabolites that may have genotoxic effects. SULT subtype 1E1 (SULT1E1) was identified as a key player in estrogen homeostasis, which is involved in many physiological processes and the pathogenesis of breast and endometrial cancer. The development of an in silico prediction model for SULT1E1 ligands would therefore support the development of metabolically inert drugs and help to assess health risks related to hormonal imbalances. Here, we report on a novel approach to develop a model that enables prediction of substrates and inhibitors of SULT1E1. Molecular dynamics simulations were performed to investigate enzyme flexibility and sample protein conformations. Pharmacophores were developed that served as a cornerstone of the model, and machine learning techniques were applied for prediction refinement. The prediction model was used to screen the DrugBank (a database of experimental and approved drugs): 28% of the predicted hits were reported in literature as ligands of SULT1E1. From the remaining hits, a selection of nine molecules was subjected to biochemical assay validation and experimental results were in accordance with the in silico prediction of SULT1E1 inhibitors and substrates, thus affirming our prediction hypotheses.Metabolism plays an important role in drug discovery and appropriate metabolic profiles of new drug candidates should be taken into consideration during the process of drug development. Acting in phase I metabolism, the enzyme family of cytochrome P450 is estimated to transform about 75% of marketed drugs (1) and numerous in silico approaches for the prediction of cytochrome P450-mediated metabolism have emerged to date (2, 3). Although the majority of metabolism prediction studies focuses on phase I, the significance of phase II metabolism is generally underestimated (4) and to this day, computer-based models for the prediction of phase II metabolism remain scarce (5).Among the predominant phase II enzyme families are the soluble sulfotransferases that form a gene superfamily termed SULT. These enzymes regulate the sulfonation of smaller molecules such as endogenous hormones, neurotransmitters, and xenobiotic substances from pharmaceutical, nutritional, or environmental sources. Based on sequence similarity, functional human SULTs are divided into two main families (SULT1 and SULT2) and further into subfamilies that exhibit individual, but somewhat overlapping substrate specificities (6). Influencing the level of female sex hormones (estrogens), SULT subtype 1E1 (SULT1E1) shows a specific substrate preference for physiological estrogenic compounds (K m ϭ 5 nM for estradiol (7)) and has been extensively investigated in experimental studies.In general, sulfonation reactions in which a sulfona...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

In Silico Prediction of Human Sulfotransferase 1E1 Activity Guided by Pharmacophores from Molecular Dynamics Simulations

Rakers

Schumacher

Meinl

et al. 2016

Journal of Biological Chemistry

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“…SULTs harbor a conserved 30-residue active-site cap that is positioned over both the nucleotide-and acceptor-binding pockets. Nucleotide binding stabilizes the cap in a "closed" position that encapsulates the nucleotide and acceptor and forms a pore that sterically restricts access to the acceptor-binding site (Cook et al, 2013c;Leyh et al, 2013;Wang et al, 2014). The cap can isomerize into an open state when nucleotide is bound, and it is from the open position that nucleotide escapes.…”

Section: Sult1a1 Allosteric Binding Sites 419mentioning

confidence: 99%

The Allosteric Binding Sites of Sulfotransferase 1A1

et al. 2014

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Human sulfotransferases (SULTs) comprise a small, 13-member enzyme family that regulates the activities of thousands of compoundsendogenous metabolites, drugs, and other xenobiotics. SULTs transfer the sulfuryl-moiety (-SO 3 ) from a nucleotide donor, PAPS (39-phosphoadenosine 59-phosphosulfate), to the hydroxyls and primary amines of acceptors. SULT1A1, a progenitor of the family, has evolved to sulfonate compounds that are remarkably structurally diverse. SULT1A1, which is found in many tissues, is the predominant SULT in liver, where it is a major component of phase II metabolism. Early work demonstrated that catechins and nonsteroidal antiinflammatory drugs inhibit SULT1A1 and suggested that the inhibition was not competitive versus substrates. Here, the mechanism of inhibition of a single, high affinity representative from each class [epigallocatechin gallate (EGCG) and mefenamic acid] is determined using initial-rate and equilibrium-binding studies. The findings reveal that the inhibitors bind at sites separate from those of substrates, and at saturation turnover of the enzyme is reduced to a nonzero value. Further, the EGCG inhibition patterns suggest a molecular explanation for its isozyme specificity. Remarkably, the inhibitors bind at sites that are separate from one another, and binding at one site does not affect affinity at the other. For the first time, it is clear that SULT1A1 is allosterically regulated, and that it contains at least two, functionally distinct allosteric sites, each of which responds to a different class of compounds.

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“…During initialrate turnover under conditions where inhibition is negligible, PAP release from the binary complex (E•PAP) is largely rate-determining; thus, E•PAP accumulates in the presteady-state phase of the reaction, contributing to a burst of product, and it reaches a fixed concentration in the steady-state. As the concentration of substrate increases, it adds more quickly to E•PAP, increasing the concentration of the dead-end complex (E•PAP•S), from which PAP escapes slowly relative to the binary complex (Wang et al, 2014a). Studies with oxidized rat enzyme implicate PAP release as the rate-liming step during uninhibited turnover of SULT1A1 (Marshall et al, 2000).…”

Section: Discussionmentioning

confidence: 99%

“…The SULTs typically exhibit partial substrate inhibition, a type of inhibition in which turnover decreases to a nonzero value at saturating substrate. Several mechanisms have been proposed to explain this inhibition in SULTs, including an allostericbinding pocket (Zhang et al, 1998), gating (Lu et al, 2008;Cook et al, 2010), the binding of multiple acceptors in the active site (Gamage et al, 2003), and the formation of a dead-end complex (Gamage et al, 2005;Tyapochkin et al, 2009;Gulcan and Duffel, 2011;Wang et al, 2014a). In a recent study of SULT2A1, each of the 22 microscopic rate constants associated with interconversion of the 11 complexes in the mechanism were determined (Wang et al, 2014a).…”

Section: Discussionmentioning

confidence: 99%

Design and Interpretation of Human Sulfotransferase A1 Assays

Wang

Cook

Leyh

2015

Drug Metabolism and Disposition

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The human sulfotransferases (SULTs) regulate the activities of hundreds, if not thousands, of small molecule metabolites via transfer of the sulfuryl-moiety (-SO 3 ) from the nucleotide donor, 39-phosphoadenosine 59-phosphosulfate (PAPS) to the hydroxyls and amines of the recipients. Our understanding of the molecular basis of SULT catalysis has expanded considerably in recent years. The basic kinetic mechanism of these enzymes, previously thought to be ordered, has been redefined as random for SULT2A1, a representative member of the superfamily. An active-site cap whose structure and dynamics are highly responsive to nucleotides was discovered and shown to be critical in determining SULT selectivity, a topic of longstanding interest to the field. We now realize that a given SULT can operate in two specificity modes-broad and narrow-depending on the disposition of the cap. More recent work has revealed that the caps of the SULT1A1 are controlled by homotropic allosteric interactions between PAPS molecules bound at the dimer's active sites. These interactions cause the catalytic efficiency of SULT1A1 to vary in a substrate-dependent fashion by as much as two orders of magnitude over a range of PAPS concentrations that spans those found in human tissues. SULT catalysis is further complicated by the fact that these enzymes are frequently inhibited by their substrates. This review provides an overview of the mechanistic features of SULT1A1 that are important for the design and interpretation of SULT1A1 assays.

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Paradigms of Sulfotransferase Catalysis

Cited by 39 publications

References 52 publications

In Silico Prediction of Human Sulfotransferase 1E1 Activity Guided by Pharmacophores from Molecular Dynamics Simulations

In Silico Prediction of Human Sulfotransferase 1E1 Activity Guided by Pharmacophores from Molecular Dynamics Simulations

The Allosteric Binding Sites of Sulfotransferase 1A1

Design and Interpretation of Human Sulfotransferase A1 Assays

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