Kinetic Mechanism and Identification of the Active Site Tyrosine Residue in Enterobacter amnigenus Arylsulfate Sulfotransferase

Kwon, Ae‐Ran; Yun, Hee-Jeong; Choi, Eung‐Chil

doi:10.1006/bbrc.2001.5185

Cited by 14 publications

(27 citation statements)

References 21 publications

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“…In contrast, the activity of the point variants Tyr208Phe and Tyr559Phe and of the double variant Tyr208Phe/Tyr559Phe was comparable to that of the wild-type variant, indicating that these tyrosines do not participate in catalysis. Similarly, replacement of Tyr-96, a previously proposed catalytic ASST residue (13), had no effect on activity (Table S2). These data do not support the previously proposed model of sulfuryl transfer from a sulfo-His to a Tyr residue in ASST before sulfurylation of the acceptor substrate, instead favoring a mechanism in which a sulfurylated activesite histidine directly delivers the sulfuryl group to the aromatic hydroxyl group of an acceptor substrate.…”

Section: Active Site Residues and The Mechanism Of Paps-independent Smentioning

confidence: 79%

“…In common with other enzymes exhibiting a ␤-propeller fold, ASST's circular array of propeller blades forms a conical central channel that is well-suited to sequester either the donor or the acceptor substrate and contains the active site of the enzyme. Previous mechanistic studies of ASST suggested the catalysis to proceed by transient tyrosine sulfurylation (1,13,31). The residue Tyr-96 was proposed to undergo sulfurylation in ASST from Enterobacter amnigenus, which is 84% identical in sequence to the ASST of E. coli CFT073 (13,31).…”

Section: Discussionmentioning

confidence: 99%

“…Previous mechanistic studies of ASST suggested the catalysis to proceed by transient tyrosine sulfurylation (1,13,31). The residue Tyr-96 was proposed to undergo sulfurylation in ASST from Enterobacter amnigenus, which is 84% identical in sequence to the ASST of E. coli CFT073 (13,31). The crystal structure presented here shows Tyr-96 to be located within the N-terminal ␤-sandwich domain far from the center of the ␤-propeller (Fig.…”

Section: Discussionmentioning

confidence: 99%

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A structural and biochemical basis for PAPS-independent sulfuryl transfer by aryl sulfotransferase from uropathogenic Escherichia coli

Malojcic

Owen

Grimshaw

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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Sulfotransferases are a versatile class of enzymes involved in numerous physiological processes. In mammals, adenosine 3-phosphate-5-phosphosulfate (PAPS) is the universal sulfuryl donor, and PAPS-dependent sulfurylation of small molecules, including hormones, sugars, and antibiotics, is a critical step in hepatic detoxification and extracellular signaling. In contrast, little is known about sulfotransferases in bacteria, which make use of sulfurylated molecules as mediators of cell-cell interactions and host-pathogen interactions. Bacterial arylsulfate sulfotransferases (also termed aryl sulfotransferases), in contrast to PAPS-dependent sulfotransferases, transfer sulfuryl groups exclusively among phenolic compounds in a PAPS-independent manner. Here, we report the crystal structure of the virulence factor arylsulfate sulfotransferase (ASST) from the prototypic, pyelonephritogenic Escherichia coli strain CFT073 at 2.0-Å resolution, and 2 catalytic intermediates, at 2.1-Å and 2.4-Å resolution, with substrates bound in the active site. ASST is one of the largest periplasmic enzymes and its 3D structure differs fundamentally from all other structurally characterized sulfotransferases. Each 63.8-kDa subunit of the ASST homodimer comprises a 6-bladed ␤-propeller domain and a C-terminal ␤-sandwich domain. The active sites of the dimer are situated at the center of the channel formed by each ␤-propeller and are defined by the side chains of His-252, His-356, Arg-374, and His-436. We show that ASST follows a ping-pong bi-bi reaction mechanism, in which the catalytic residue His-436 undergoes transient sulfurylation, a previously unreported covalent protein modification. The data provide a framework for understanding PAPS-independent sulfotransfer and a basis for drug design targeting this bacterial virulence factor. beta propeller ͉ crystal structure ͉ pyelonephritis ͉ uropathogenic Escherichia coli CFT073 periplasm S ulfotransferases catalyze the transfer of a sulfuryl group from an activated donor to an acceptor and are essential for numerous physiological processes, such as sulfur metabolism, liver detoxification, signal transduction, hormone regulation, viral entry, and molecular recognition. A variety of small molecules, including hormones, sugars, and antibiotics, have been shown to undergo 3Ј-phosphate-5Ј-phosphosulfate (PAPS)-dependent sulfurylation (1, 2), and sulfotransferases are recognized as modulators of prokaryote-eukaryote interactions (3).In mammals, sulfoconjugation and glucuronidation represent the dominant mechanisms for detoxification of endogenous and exogenous compounds bearing phenolic groups (4, 5). Therefore, hepatic sulfotransferases are of considerable toxicological and pharmacological interest. In addition to mammalian sulfotransferases, arylsulfate sulfotransferases of commensal intestinal bacteria have been proposed to play a role in the detoxification of phenolic compounds (6, 7). Although a number of eukaryotic sulfotransferases have been extensively studied, much less is known about ba...

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Section: Active Site Residues and The Mechanism Of Paps-independent Smentioning

confidence: 79%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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A structural and biochemical basis for PAPS-independent sulfuryl transfer by aryl sulfotransferase from uropathogenic Escherichia coli

Malojcic

Owen

Grimshaw

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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“…ASST activity was quantified by monitoring the production of pnitrophenol from the hydrolysis of p-nitrophenylsulfate (PNS), using a-naphthol as an acceptor (Kwon et al, 2001). Cultures were grown overnight at 37 uC in LB broth with 2 mM DTT and 100 mg ampicillin ml 21 .…”

Section: Methodsmentioning

confidence: 99%

DsbL and DsbI contribute to periplasmic disulfide bond formation in Salmonella enterica serovar Typhimurium

2009

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Disulfide bond formation in periplasmic proteins is catalysed by the DsbA/DsbB system in most Gram-negative bacteria. Salmonella enterica serovar Typhimurium also encodes a paralogous pair of proteins to DsbA and DsbB, DsbL and DsbI, respectively, downstream of a periplasmic arylsulfate sulfotransferase (ASST). We show that DsbL and DsbI function as a redox pair contributing to periplasmic disulfide bond formation and, as such, affect transcription of the Salmonella pathogenicity island 1 (SPI1) type three secretion system genes and activation of the RcsCDB system, as well as ASST activity. In contrast to DsbA/DsbB, however, the DsbL/DsbI system cannot catalyse the disulfide bond formation required for flagellar assembly. Phylogenic analysis suggests that the assT dsbL dsbI genes are ancestral in the Enterobacteriaceae, but have been lost in many lineages. Deletion of assT confers no virulence defect during acute Salmonella infection of mice.

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“…[68] Investigations with the aryl sulfotransferase from Enterobacter amnigenus led to the same characteristic double reciprocal pattern, as well as to the finding that PNP is a noncompetitive inhibitor of the sulfate donor PNPS and a competitive inhibitor of the acceptor 1-naphthol. [69] 5.3. Transition-State Structure…”

Section: Sulfotransferasesmentioning

confidence: 99%

Sulfotransferases: Structure, Mechanism, Biological Activity, Inhibition, and Synthetic Utility

et al. 2004

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The sulfonation (also known as sulfurylation) of biomolecules has long been known to take place in a variety of organisms, from prokaryotes to multicellular species, and new biological functions continue to be uncovered in connection with this important transformation. Early studies of sulfotransferases (STs), the enzymes that catalyze sulfonation, focused primarily on the cytosolic STs, which are involved in detoxification, hormone regulation, and drug metabolism. Although known to exist, the membrane-associated STs were not studied as extensively until more recently. Involved in the sulfonation of complex carbohydrates and proteins, they have emerged as central players in a number of molecular-recognition events and biochemical signaling pathways. STs have also been implicated in many pathophysiological processes. As a result, much interest in the complex roles of STs and in their targeting for therapeutic intervention has been generated. Progress in the elucidation of the structures and mechanisms of sulfotransferases, as well as their biological activity, inhibition, and synthetic utility, are discussed in this Review.

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Kinetic Mechanism and Identification of the Active Site Tyrosine Residue in Enterobacter amnigenus Arylsulfate Sulfotransferase

Cited by 14 publications

References 21 publications

A structural and biochemical basis for PAPS-independent sulfuryl transfer by aryl sulfotransferase from uropathogenic Escherichia coli

A structural and biochemical basis for PAPS-independent sulfuryl transfer by aryl sulfotransferase from uropathogenic Escherichia coli

DsbL and DsbI contribute to periplasmic disulfide bond formation in Salmonella enterica serovar Typhimurium

Sulfotransferases: Structure, Mechanism, Biological Activity, Inhibition, and Synthetic Utility

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