Mechanistic studies of β-arylsulfotransferase IV

Abstract

Sulfotransferases are an important class of enzymes that catalyze the transfer of a sulfuryl group to a hydroxyl or amine moiety on various molecules including small-molecule drugs, steroids, hormones, carbohydrates, and proteins. They have been implicated in a number of disease states but remain poorly understood, complicating the design of specific, small-molecule inhibitors. A linear free-energy analysis in both the forward and reverse directions indicates that the transfer of a sulfuryl group to an aryl hydroxyl group catalyzed by ␤-arylsulfotransferase IV likely proceeds by a dissociative (sulfotrioxide-like) mechanism. Values for the Brønsted coefficients (␤ nuc and ␤lg) are ؉0.33 and ؊0.45, giving Leffler ␣ values of 0.19 and 0.61 for the forward and reverse reactions, respectively.sulfotransferase ͉ linear free energy ͉ mechanism S ulfotransferases (STs) are an important class of enzymes that catalyze the transfer of a sulfuryl group from 3Ј-phospho-adenosyl-5Ј-phosphosulfate (PAPS) to a hydroxyl or, less frequently, an amine to give the sulfated product and the cofactor byproduct 3Ј-phosphoadenosyl-5Ј-phosphate (PAP) (Fig. 1). They can be divided into two groups: the cytosolic STs and the membrane-associated STs. Cytosolic STs catalyze the sulfonation of xenobiotics for detoxification and hormones for regulation. Membrane-associated STs catalyze the sulfonation of proteins and carbohydrates for processes such as cellular signaling and modulation of receptor binding (1-9). Recent studies have implicated the STs in a number of disease states including entry of the herpes virus (10), entry of HIV (11, 12), chronic inf lammation (13), and various forms of cancer (1-9). For this reason there is considerable interest in the design of small-molecule inhibitors that are specific for a particular ST. Although a number of inhibitor designs and syntheses have been published, to date the inhibitors are relatively weak and have always been discovered through semirational methods because of the lack of a complete mechanistic picture (14).A number of structural and mechanistic studies have been carried out on STs. Crystal structures have been solved for estrogen ST (15), catecholamine ST (16), dopamine ST (17), hydroxysteroid ST (18), and the ST domain of heparan sulfate N-deacetylase͞N- . Sequence alignment data from these structures as well as other STs show there to be a great deal of homology at the amino acid level for certain residues near the active site. Furthermore, mutagenesis studies carried out on these conserved residues show several of them to be critical to enzyme function. One such study of estrogen ST using a bound vanadate indicates an in-line attack of PAPS by the steroid hydroxyl group (20). Moreover, early studies of ␤-arylsulfotransferase IV (␤-AST-IV) show a random, rapidly equilibrating bi-bi mechanism (21). In the kinase field, the transition state of a protein tyrosine kinase has been illuminated through the use of linear free-energy studies (22). This information then was used to design spec...

Physicochemical properties of hydroxylated polychlorinated biphenyls aid in predicting their interactions with rat sulfotransferase 1A1 (rSULT1A1)

“…These findings with the OHPCBs are consistent with a previous report on fluoronitrophenols as substrates for rSULT1A1, in which the log of the apparent second-order rate constant (K cat /K m ) of each compound paralleled the corresponding pKa value [50]. Moreover, our examination of the relationship between the pKa values for OHPCBs and their log IC 50 values for inhibition of rSULT1A1 shown in Figure 2 revealed a relationship wherein the most potent inhibitors (i.e., those with IC 50 values lower than 4.4 µM) had lower pKa values, while the less potent inhibitors (i.e., those with IC 50 values greater than 26 µM) had higher pKa values.…”

“…This would be consistent with structural studies on the mechanism of a sulfotransferase-catalyzed reaction where the catalytic histidine is involved in orientation of a substrate and abstraction of the phenolic proton during an in-line sulfuryl transfer [51]. Likewise, this is consistent with linear free-energy analyses [50] and kinetic isotope studies [52] that indicate a relatively loose, dissociative transition state in the transfer of the sulfuryl group in the mechanisms of sulfotransferases. Those OHPCBs with very low pKa values, such as the 4-OH-3,5-chloro-substituted PCBs, bind to the enzyme well (as seen by their log K d values), however, they may not be in the correct orientation for sulfuryl transfer to occur.…”

Spectroscopic Studies on the [4Fe-4S] Cluster in Adenosine 5′-Phosphosulfate Reductase from Mycobacterium tuberculosis

“…Although only speculative at present, it is plausible that the active site Lys-144 cation may orient the incoming Cys nucleophile with respect to the sulfate moiety and/or act as a "molecular guide wire" during sulfuryl transfer; either possibility could result in transition state stabilization. Analogous functions for Lys and Arg residues in the active site of other enzymes that catalyze sulfuryl transfer have been described (55,56).…”

Effiziente katalytische Promiskuität für chemisch unterschiedliche Reaktionen

“…Die tatsächlichen Charakteristika der Übergangszustände in der enzymatischen Reaktion sind noch zu bestimmen. AP, [27,28] einige andere Phosphatasen [22,29] und eine Sulfatase [30] nutzen Übergangs-zustände, die auch bei der unkatalysierten Reaktion in freier Lösung dominieren. Allerdings gibt es auch Gegenbeispiele, z.…”