The Pharmacogenomics Journal (2002) 2, 297–308 & 2002 Nature Publishing Group All rights reserved 1470-269X/02 $25.00 www.nature.com/tpj REVIEW Sulfation through the looking glass—recent advances in sulfotransferase research for the curious MWH Coughtrie ABSTRACT Members of the cytosolic sulfotransferase (SULT) superfamily catalyse the Department of Molecular & Cellular Pathology, sulfation of a multitude of xenobiotics, hormones and neurotransmitters. University of Dundee, Ninewells Hospital & Humans have at least 10 functional SULT genes, and a number of recent Medical School, Dundee, Scotland, UK advances reviewed here have furthered our understanding of SULT function. Correspondence: Analysis of expression patterns has shown that sulfotransferases are highly MWH Coughtrie, Department of expressed in the fetus, and SULTs may in fact be a major detoxification Molecular & Cellular Pathology, University enzyme system in the developing human. The X-ray crystal structures of of Dundee, Ninewells Hospital and three SULTs have been solved and combined with mutagenesis experiments Medical School, Dundee DD1 9SY, Scotland, UK. and molecular modelling, they have provided the first clues as to the factors Tel: +44 (0)1382 632510 that govern the unique substrate specificities of some of these enzymes. In Fax: +44 (0)1382 640320 the future these and other studies will facilitate prediction of the fate of E-mail: [email protected] chemicals metabolised by sulfation. Variation in sulfation capacity may be important in determining an individual’s response to xenobiotics, and there has been an explosion in information on sulfotransferase polymorphisms and their functional consequences, including the influence of SULT1A1 genotype on susceptibility to colorectal and breast cancer. Finally, the first gene knockout experiments with SULTs have recently been described, with the generation of estrogen sulfotransferase deficient mice in which reproductive capacity is compromised. Our improved understanding of these enzymes will have significant benefits in such diverse areas as drug design and develop- ment, cancer susceptibility, reproduction and development. The Pharmacogenomics Journal (2002) 2, 297–308. doi:10.1038/sj.tpj.6500117 Keywords: sulfotransferase; sulfation; pharmacogenetics; structural biology; development Sulfation of a vast array of natural and synthetic chemicals as well as many biomolecules occurs widely in nature, from bacteria to humans. This article concerns the formation of sulfate conjugates of xenobiotic and endogenous (endobiotic) small molecules by members of the cytosolic sulfotransferase (SULT) enzyme family. The process was first discovered in the late 1870s by Eugen Baumann, who isolated and characterised the sulfate conjugate of phenol from urine of a patient treated with carbolic acid as an antiseptic. It would be another 80 years, however, before the mechanistic basis of sulfate conjugation could be identified, with the discovery of so-called ‘active sulfate’ by Lipmann’s group.1 This compound, that we now call PAPS (30-phosphoadenosine 50-phosphosul- fate), is the sulfuryl donor for the vast majority of sulfation reactions2 (Box 1). Recent research in this field that will be highlighted here, has uncovered some important aspects of the function of sulfation. The relative importance of sulfation in humans has perhaps been underestimated in the past, and one of the main reasons for this is that there are major inter-species differences in the function of sulfation—in particular relating to its role in modulating the function of Received: 28 February 2002 Revised: 28 March 2002 hormones and neurotransmitters. Importantly, catecholamines, estrogens, Accepted: 4 April 2002 iodothyronines and dehydroepiandrosterone (DHEA) exist in the human Recent advances in sulfotransferase research MWH Coughtrie 298 Box 1 - Sulfotransferase Reaction and Co-substrate Synthesis Paracetamol Paracetamol Sulfate NH CH NH CH3 3 Sulfotransferase O O HO HO 3S 2 APS PAPS PAP ATP 1 ATP ADP PPi SO4-- The sulfotransferase reaction relies on the co-substrate PAPS as sulfuryl donor. PAPS is synthesised from inorganic sulfate and 2 molecules of ATP - it is therefore an "expensive" commodity for cells to produce, and is present at relatively low concentrations, typically no more than 20 or 30µM. Low PAPS levels and rapid depletion is believed to limit the capacity of sulfation compared to e.g. glucuronidation, where the co-substrate (UDP-glucuronic acid) is in more plentiful supply. In animals, synthesis of PAPS is the responsibility of a single bifunctional enzyme called PAPS synthetase (PAPSS) that contains both the ATP sulfurylase (1) and APS kinase (2) catalytic activities. In humans and mice, 2 isoforms of PAPSS exist that are the products of the PAPSS1 and PAPSS2 genes. Mutations in PAPSS2 are responsible for rare inherited connective tissue disorders in mouse (brachymorphism) and humans (spondyloepimetaphyseal dysplasia). It is postulated that interindividual variation in the ability to synthesise PAPS, arising from genetic and/or environmental effects on PAPSS, may influence sulfation capacity. circulation predominantly or significantly as the sulfated SULTs related to the mammalian cytosolic enzymes, that are form, whereas this is not the case for rodents or many other involved in flavonol and brassinosteroid metabolism.16 ‘experimental’ animals.3 This is reflected in the different Other model organisms whose genomes have been (or are SULT isoenzyme profiles that exist in various species. For being) sequenced possess various SULTs—the zebrafish example, to date human is the only species in which a (Danio rerio) and amphibian (Xenopus laevis, Silurana tropi- catecholamine-specific SULT isoform (SULT1A3) has been calis) sequence databases contain numerous cytosolic SULT identified, consistent with the high circulating levels of sequences with orthologs in mammalian species, although catecholamine sulfates found in humans and other pri- the Drosophila melanogaster, Caenorhabditis elegans and mates.3,4 Rodents have multiple isoforms of the hydroxyster- Saccharomyces cerevisiae databanks are almost devoid of such oid SULT subfamily 2A,5–7 whereas so far only a single sequences. A new nomenclature system for the cytosolic SULT2A enzyme has been identified and characterised in SULTs has recently been developed* which will (hopefully) humans.8 Also, it is now clear that SULTs are abundantly help to clarify the relationships between the various SULTs expressed in the human fetus9–13—other ‘drug metabolising for novices and aficionados alike. The nomenclature used enzymes’ such as the cytochromes P450 (CYPs), UDP- here follows this new system. glucuronosyltransferases (UGTs) etc are, in general not The SULT1 and SULT2 families are the largest, and expressed at significant levels until after birth14,15—thus probably the most important for xenobiotic and endobiotic SULTs may represent a front line of chemical defence in the metabolism. At present the human SULT enzyme family, developing human. which is the best characterised, comprises of 11 isoforms This article aims to provide the reader with an up-to-date representing the SULT1, SULT2 and SULT4 families. These overview of sulfation in humans, and to highlight some enzymes are the products of 10 genes (SULTs 2B1a and 2B1b important recent research that has impacted significantly on are generated by alternate splicing of the first exon of our knowledge and understanding of this important meta- SULT2B1) that share many common structural features bolic system. (reviewed in17). The properties of the various human SULTs are summarised in Table 1. The results of many studies show THE SULFOTRANSFERASE SUPERFAMILY— clearly that the expression of SULTs in humans is carefully ORGANISATION AND FUNCTION regulated with respect to tissue type, development and As with many drug metabolising enzymes, the cytosolic hormonal influences, supporting proposed functional roles SULTs are derived from a large superfamily of genes. Full- for some of the enzymes. A number of examples are worth length cDNAs encoding more than 50 mammalian and avian cytosolic SULTs have been cloned and sequenced, and *Raftogianis RB et al, submitted for publication. In particular, the many of the expressed proteins characterised. The majority new nomenclature for the SULT1C enzymes may cause some of these enzymes sulfate small molecule xenobiotics and/or confusion. The human SULT1C2 referred to here was called endogenous hormones and neurotransmitters, and based on SULT1C1 or SULT1C sulfotransferase 1 in the original descrip- amino acid sequence analysis can be subdivided into six tions,18,19,75 and the human SULT1C4 referred to here was originally families (Box 2). In addition plants possess a number of called SULT1C2 or SULT1C sulfotransferase.19,75 The Pharmacogenomics Journal Recent advances in sulfotransferase research MWH Coughtrie 299 considering further: SULTs 1C2 and 1C4 appear to be most 17b-estradiol, suggesting an important role for this enzyme highly expressed in fetal tissues,13 although RNA dot blots in modulating estrogen action.21 Indeed, the enzyme is indicated the adult stomach and kidney (SULT1C2) and expressed in the endometrium and is exquisitely regulated ovary (SULT1C4) may be sites of expression.18,19 However during the menstrual cycle22,23 (probably under the primary the function of these enzymes in humans is not clear. The influence of progesterone24,25) where it is postulated to catecholamine sulfotransferase