title: Variants of glutathione s-transferase pi 1 exhibit differential enzymatic activity and inhibition by heavy metals.
abstract: Nonsynonymous single nucleotide polymorphisms in glutathione s-transferase pi 1 (GSTP1; Ile/Val 105, Ala/Val 114) have been associated with altered toxicant metabolism in epidemiological cohorts. We explored the impact of GSTP1 genotype on enzyme kinetics and heavy metal inhibition in vitro. Four GSTP1 allozymes (105/114: Ile/Ala, Val/Ala, Ile/Val, Val/Val) were expressed in and purified from Escherichia coli. Enzyme activity assays quantifying the rate of glutathione conjugation with 1-chloro-2,4-dinitrobenzene (CDNB) revealed significant differences in kinetic parameters depending on genotype (p<0.01). Allozymes with Ile105 had better catalytic efficiency and greater affinity for CDNB (mean ± SEM: Ile105 Ala114 K(m)=0.33 ± 0.07 mM vs. Val105 Ala114 K(m)=1.15 ± 0.07 mM). Inhibition of GSTP1 activity by heavy metals was assessed following treatment with mercury (inorganic-HgCl(2), methylmercury-MeHg), selenium, cadmium, lead, arsenic, and manganese. All allozymes were inhibited by HgCl(2) (IC(50) range: 24.1-172 μM), MeHg (93.9-480 μM), and selenium (43.7-62.8 μM). Genotype significantly influenced the potency of mercury with GSTP1 Ile105 Val114 the least sensitive and Val105 Ala114 the most sensitive to inhibition by HgCl(2) and MeHg. Overall, genotype of two nonsynonymous polymorphisms in GSTP1 influenced enzyme kinetics pertaining to an electrophilic substrate and inhibition by two mercury species.
authors.affiliation: Department of Environmental Health Sciences, University of Michigan School of Public Health, 1415 Washington Heights, Ann Arbor, MI 48109, USA. gaydojac@umich.edu
authors.forename: Jaclyn M
authors.initials: JM
authors.lastname: Goodrich
authors.forename: Niladri
authors.initials: N
authors.lastname: Basu
mesh_terms: D001151:Arsenic / Q000633:toxicity*; D004137:Dinitrochlorobenzene / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005838:Genotype; D005978:Glutathione / Q000378:metabolism; D051549:Glutathione S-Transferase pi / Q000235:genetics / Q000378:metabolism*; D007527:Isoenzymes; D019216:Metals, Heavy / Q000633:toxicity*; D008767:Methylmercury Compounds / Q000633:toxicity*; D009154:Mutation; D012643:Selenium / Q000633:toxicity*
chemical_list: D004137:Dinitrochlorobenzene; D007527:Isoenzymes; D019216:Metals, Heavy; D008767:Methylmercury Compounds; C496556:GSTP1 protein, human; D051549:Glutathione S-Transferase pi; D005978:Glutathione; D012643:Selenium; D001151:Arsenic