Expression, purification and characterization of human cytosolic sulfotransferase (SULT) 1C4

Abstract

Human cytosolic sulfotransferase 1C4 (hSULT1C4) is a dimeric Phase II drug-metabolizing enzyme primarily expressed in the developing fetus. SULTs facilitate the transfer of a hydrophilic sulfonate moiety from 3′-phosphoadenosine-5′-phosphosulfate (PAPS) onto an acceptor substrate altering the substrate’s biological activity and increasing the compound’s water solubility. While several of the hSULTs’ endogenous and xenobiotic substrates have been identified, the physiological function of hSULT1C4 remains unknown. The fetal expression of hSULT1C4 leads to the hypothesis that the function of this enzyme may be to regulate metabolic and hormonal signaling molecules, such as estrogenic compounds, that may be generated or consumed by the mother during fetal development. Human SULT1C4 has previously been shown to sulfonate estrogenic compounds, such as catechol estrogens; therefore, this study focused on the expression and purification of hSULT1C4 in order to further characterize this enzyme’s sulfonation of estrogenic compounds. Molecular modeling of the enzyme’s native properties helped to establish a novel purification protocol for hSULT1C4. The optimal activity assay conditions for hSULT1C4 were determined to be pH 7.4 at 37°C for up to 10 min. Kinetic analysis revealed the enzyme’s reduced affinity for PAPS compared to PAP. Human SULT1C4 sulfonated all the estrogenic compounds tested, including dietary flavonoids and environmental estrogens; however, the enzyme has a higher affinity for sulfonation of flavonoids. These results suggest hSULT1C4 could be metabolizing and regulating hormone signaling pathways during human fetal development.

Bioaccessibility and bioavailability of phenolic compounds

“…Herein, genistein-7-glucoside has a higher affinity for β-glucosidase than quercetin-4′-glucoside thus the former shows a higher deglycosylation rate, whereas some other glycosides such as quercetin-3,4′-diglucoside, quercetin-3-glucoside, kaempferol-3-glucoside, quercetin-3-rhamnoglucoside, and naringenin-7-rhamnoglucoside prefer to remain unchanged (Day et al, 1998). This intracellular reaction is regarded as a vital way to expose hydroxyl group of xenobiotics for further conjugation Cheng et al (1999), Margaillan et al (2015), Wong et al (2009), Finel et al (2005), King et al (1999), Mostaghel et al (2016), King et al (2000), Teubner et al (2007), Cheng et al (1998), Cheng et al (1999), Barbier et al (2000), Knights et al (2013), Chimalakonda et al (2011), Riches et al (2009), Ghosh et al (2013), Sakamoto et al (2015), Lu et al (2015), Guidry et al (2017), Kurogi et al (2017),…”

Regulation of Cytosolic Sulfotransferases in Models of Human Hepatocyte Development

“…These data show that SULT1A1, SULT1C2, SULT1E1, and SULT2A1 mRNA and protein are substantially expressed in prenatal liver, whereas SULT1C4 is substantially expressed only at the mRNA level. SULT1C4 was recently shown to metabolize estrogenic compounds, including estradiol and environmental estrogens (Guidry et al, 2017). Although hepatic expression of the SULT1C genes and SULT1E1 decreases during the transition from fetal to adult life, SULT1A1 expression stays relatively constant whereas SULT2A1 expression increases.…”

Developmental Expression of SULT1C4 Transcript Variants in Human Liver: Implications for Discordance Between SULT1C4 mRNA and Protein Levels

“…SULT1C4 is a member of a gene subfamily that includes three human members, SULT1C2,1C3, and 1C4. Several studies have suggested that SULT1C4 has the highest sulfonation capacity of the SULT1C enzymes towards xenobiotics, including a wide range of drugs (e.g., acetaminophen), environmental chemicals (e.g., bisphenol A), and procarcinogens (e.g., hydroxymethyl furans) (Sakakibara et al, 1998;Glatt et al, 2004;Allali-Hassani et al, 2007;Yasuda et al, 2007;Glatt et al, 2012;Yamamoto et al, 2015;Yamamoto et al, 2016;Guidry et al, 2017;Rasool et al, 2017). SULT1C4 is also capable of metabolizing estrogenic compounds, such as catechol and methoxy estrogens (Allali-Hassani et al, 2007;Hui et al, 2008).…”

“…SULT1C4 is also capable of metabolizing estrogenic compounds, such as catechol and methoxy estrogens (Allali-Hassani et al, 2007;Hui et al, 2008). Guidry et al (Guidry et al, 2017) recently reported that SULT1C4 has high sulfonation capacity towards dietary flavonoids and environmental estrogens.…”

“…Expression of SULT1C4 TVs in HEK293T cells. 5'-Asp-Tyr-Lys-Asp-Asp-Asp-Asp-Lys (DDK)tagged SULT1C4 TV1 coding sequence was prepared using 100 ng of pKK233-2-SULT1C4 (Guidry et al, 2017) as template, Herculase II Fusion DNA Polymerase (Agilent Technologies), the SULT1C4 primer pair indicated in Supplemental Table 1 (SULT1C4-HindIII-DDK-F and SULT1C4-XhoI-R), and the following PCR conditions: 95C for 2 minutes; 20 cycles of 95C for 20 seconds, 64C for 20 seconds, and 72C for 30 seconds; and 72C for 3 minutes. DDKtagged SULT1C4 TV2 and E3DEL were prepared using the TV2 and E3DEL plasmid standards (described in the section above) as templates, HotStar Taq DNA Polymerase (Qiagen), and the same primer pair used for TV1.…”