Structure-activity studies of sulfate transfer: the hydrolysis and aminolysis of 3′-phosphoadenosine 5′-phosphosulfate (PAPS)

Bedford, Colin T.; Kirby, Anthony J.; Logan, C.; Drummond, Jeremy N.

doi:10.1016/0968-0896(95)00015-9

Cited by 15 publications

(12 citation statements)

References 8 publications

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“…Kinetic isotope effects for the hydrolysis of pNPS by the arylsulfatases from Helix pomatia and Aerobacter aerogenes found 18 (V/ K) nonbridge values similar to those of the EST reaction, indicating little involvement of the nonbridging oxygen atoms and little nucleophilic involvement (15). The uncatalyzed sulfuryl transfer from PAPS to water is believed to proceed by a sulfur trioxide-like transition state with only weak involvement of water (27), like the uncatalyzed hydrolysis reactions of aryl sulfates. Thus, the evidence available thus far, in the few enzymatic sulfuryl transfer reactions that have been studied to date, suggests that enzymatic and uncatalyzed sulfuryl transfer reactions proceed with similar transition states.…”

Section: Resultsmentioning

confidence: 95%

Transition State of the Sulfuryl Transfer Reaction of Estrogen Sulfotransferase

Hoff

Czyryca

Sun

et al. 2006

Journal of Biological Chemistry

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In 1876, Baumann (1) demonstrated that phenol administered to a patient was excreted as phenyl sulfate. It was not until some 80 years later that the most common biological sulfate donor, 3Ј-phosphoadenosine-5Ј-phosphosulfate (PAPS), 3 was identified (2). Sulfotransferases catalyze the transfer of the sulfuryl group from a donor, which is most often PAPS, to hydroxyl or amine nucleophiles (Fig. 1). The reverse reaction, the hydrolysis of sulfate esters, is catalyzed by sulfatases. Thus, these enzymes have a complementary relationship analogous to that of kinases and phosphatases. However, enzymatic sulfuryl transfer has remained less thoroughly studied than phosphoryl transfer. Among sulfotransferases that utilize PAPS, several structural and mechanistic investigations have been reported. Structures have been solved for catecholamine sulfotransferase (3), dopamine sulfotransferase (4), and hydroxysteroid sulfotransferase (5). The x-ray structures have been solved for estrogen sulfotransferase (EST) with 3Ј-phosphoadenosine 5Ј-phosphate (PAP) and estradiol bound (6) and also for the EST-PAP-vanadate complex (7).EST catalyzes sulfuryl transfer between PAPS and the phenolic oxygen atom of ␤-estradiol. The sulfation prevents ␤-estradiol from binding to and activating the estrogen receptor. EST catalyzes both the forward and the reverse reactions, as shown in Fig. 1; p-nitrophenol is an alternate sulfuryl acceptor in the forward direction, and p-nitrophenyl sulfate ( pNPS) can function as a sulfuryl donor to PAP in the reverse direction (8).X-ray structures and site-directed mutagenesis have identified two crucial active site residues, His-108 and Lys-106, which are each ϳ3 Å from the oxygen atom of estradiol. Mutation of Lys-106 to arginine, serine, and alanine had little impact on the K m for PAPS but decreased the k cat value by a factor of 200 -3000, and mutation of His-108 to serine, leucine, lysine, or arginine resulted in a protein with no detectable catalytic activity (7). This effect and the position of His-108 relative to the phenol group on the substrate suggest a role for this residue as a catalytic base. Lys-106, the only residue in a position to interact with the nonbridging oxygens of the transferring SO 3 moiety, probably functions in substrate positioning and transition state stabilization. The x-ray structures led to the proposal of a concerted, in-line mechanism for transfer of the sulfuryl group (7).Kinetic isotope effects (9) and linear free energy relationships (10 -14) have revealed that uncatalyzed sulfuryl transfer reactions have transition states characterized by extensive leaving group bond fission and little nucleophilic participation. This is described as a loose transition state in which the sulfuryl group resembles SO 3 . In this study, kinetic isotope effects were used to examine the transition state of the EST-catalyzed sulfuryl transfer from the alternate sulfuryl donor pNPS to PAP. The KIEs have previously been reported for the uncatalyzed hydrolysis of pNPS (9) and for the reaction...

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Section: Resultsmentioning

confidence: 95%

Transition State of the Sulfuryl Transfer Reaction of Estrogen Sulfotransferase

Hoff

Czyryca

Sun

et al. 2006

Journal of Biological Chemistry

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“…An alternative function would be the stabilization of the developing negative charge on the bridging oxygen of the 5′-phosphate. From studies on the chemical hydrolysis and aminolysis of PAPS, it was proposed that in the transition state, the leaving group is highly polarized, possessing two almost fully formed negative charges (28). The catalytic function of Lys-59 might be similar to that of the invariant lysine present in the P-loop of adenylate kinases, which was proposed to accompany the transferred negative charge (29).…”

Section: Discussionmentioning

confidence: 99%

3‘-Phosphoadenosine 5‘-Phosphosulfate Binding Site of Flavonol 3-Sulfotransferase Studied by Affinity Chromatography and ³¹P NMR

et al. 1999

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The function of Lys-59, Arg-141, and Arg-277 in PAPS binding and catalysis of the flavonol 3-sulfotransferase was investigated. Affinity chromatography of conservative mutants with PAPS analogues allowed us to determine that Lys-59 interacts with the 5' portion of the nucleotide, while Arg-141 interacts with the 3' portion, confirming assignments deduced from the crystal structure of mouse estrogen sulfotransferase [Kakuta, Y., Pedersen, L. G., Carter, C. W. , Negishi, M., and Pedersen, L. C. (1997) Nat. Struct. Biol. 4, 904-908]. The affinity chromatography method could be used to characterize site-directed mutants for other types of enzymes that bind nucleoside 3',5'- or 2',5'-diphosphates. 31P NMR spectra of enzyme-PAP complexes were recorded for the wild-type enzyme and K59R and K59A mutants. The results of these experiments suggest that Lys-59 is involved in the determination of the proper orientation of the phosphosulfate group for catalysis.

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“…stable, an accelerated breakdown by aminolysis might be expected (27) during crystallization, which in our experiment required incubation for several days in Tris buffer.…”

Section: Resultsmentioning

confidence: 99%

The Structure of Human SULT1A1 Crystallized with Estradiol

Gamage

Tsvetanov

Duggleby

et al. 2005

Journal of Biological Chemistry

107

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Human SULT1A1 belongs to the supergene family of sulfotransferases (SULTs) involved in the sulfonation of xeno-and endobiotics. The enzyme is also one of the SULTs responsible for metabolic activation of mutagenic and carcinogenic compounds and therefore is implicated in various cancer forms. Further, it is not well understood how substrate inhibition takes place with rigid fused multiring substrates such as 17␤-estradiol (E2) at high substrate concentrations when subcellular fractions or recombinant enzymes are used. To investigate how estradiol binds to SULT1A1, we co-crystallized SULT1A1 with sulfated estradiol and the cofactor product, PAP (3-phosphoadenosine 5-phosphate). The crystal structure of SULT1A1 that we present here has PAP and one molecule of E2 bound in a nonproductive mode in the active site. The structure reveals how the SULT1A1 binding site undergoes conformational changes to accept fused ring substrates such as steroids. In agreement with previous reports, the enzyme shows partial substrate inhibition at high concentrations of E2. A model to explain these kinetics is developed based on the formation of an enzyme⅐PAP⅐E2 dead-end complex during catalysis. This model provides a very good quantitative description of the rate versus the [E2] curve. This dead-end complex is proposed to be that described by the observed structure, where E2 is bound in a nonproductive mode. Cytosolic sulfotransferases (SULTs)2 are involved in the phase II metabolism of numerous xeno-and endobiotics such as drugs, neurotransmitters, bile acids, and hormones (1, 2). SULTs utilize the cofactor 3Ј-phosphoadenosine 5Ј-phosphosulfate (PAPS) as the sulfonate (SO 3 Ϫ1 ) donor in such reactions that generally lead to the detoxification of substrates by making them more water-soluble and thereby readily excretable via the kidneys. However, in the case of mutagenic and carcinogenic N-hydroxyarylamines and heterocyclic amines and benzylic alcohols of polycyclic aromatic hydrocarbons, sulfonation is a critical step in their metabolic activation to electrophiles that can bind tissue macromolecules such as DNA (3-6). Further, sulfonation has been shown to activate the antihypertensive and hair growth stimulant minoxidil (7). The overall role that sulfonation plays in disease states such as neurodegeneration and cancer is currently under active investigation (8 -10). SULT1A1 has a broad tissue distribution and has been shown to metabolize a wide range of xenobiotics and play a significant role in the metabolism of estrogens and iodothyronines. In resolving the crystal structure (11) of SULT1A1 we showed that it contains a very hydrophobic L-shaped substrate-binding region, which explains how it can accommodate small planar compounds as well as larger L-shaped aromatics such as iodothyronines. However, in those studies we could not explain how extended fused ring systems, such as 17␤-estradiol (E2), can act as substrates of SULT1A1. In addition, no real information has been provided as to why the metabolism of E2 is inhibited at high...

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Structure-activity studies of sulfate transfer: the hydrolysis and aminolysis of 3′-phosphoadenosine 5′-phosphosulfate (PAPS)

Cited by 15 publications

References 8 publications

Transition State of the Sulfuryl Transfer Reaction of Estrogen Sulfotransferase

Transition State of the Sulfuryl Transfer Reaction of Estrogen Sulfotransferase

3‘-Phosphoadenosine 5‘-Phosphosulfate Binding Site of Flavonol 3-Sulfotransferase Studied by Affinity Chromatography and ³¹P NMR

The Structure of Human SULT1A1 Crystallized with Estradiol

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Structure-activity studies of sulfate transfer: the hydrolysis and aminolysis of 3′-phosphoadenosine 5′-phosphosulfate (PAPS)

Cited by 15 publications

References 8 publications

Transition State of the Sulfuryl Transfer Reaction of Estrogen Sulfotransferase

Transition State of the Sulfuryl Transfer Reaction of Estrogen Sulfotransferase

3‘-Phosphoadenosine 5‘-Phosphosulfate Binding Site of Flavonol 3-Sulfotransferase Studied by Affinity Chromatography and 31P NMR

The Structure of Human SULT1A1 Crystallized with Estradiol

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3‘-Phosphoadenosine 5‘-Phosphosulfate Binding Site of Flavonol 3-Sulfotransferase Studied by Affinity Chromatography and ³¹P NMR