Abstract
Polycyclic aromatic hydrocarbons (PAH) are environmental and tobacco carcinogens. Human aldo-keto reductases catalyze the metabolic activation of proximate carcinogenic PAH trans-dihydrodiols to yield electrophilic and redox-active o-quinones. Benzo[a]pyrene-7,8-dione a representative PAH o-quinone is reduced back to the corresponding catechol to generate a futile redox-cycle. We investigated whether sulfonation of PAH catechols by human sulfotransferases (SULT) could intercept the catechol in human lung cells. RT-PCR identified SULT1A1, -1A3, and -1E1 as the isozymes expressed in four human lung cell lines. The corresponding recombinant SULTs were examined for their substrate specificity. Benzo[a]pyrene-7,8-dione was reduced to benzo[a]pyrene-7,8-catechol by dithiothreitol under anaerobic conditions and then further sulfonated by the SULTs in the presence of 3'-[(35)S]phosphoadenosine 5'-phosphosulfate as the sulfonate group donor. The human SULTs catalyzed the sulfonation of benzo[a]pyrene-7,8-catechol and generated two isomeric benzo[a]pyrene-7,8-catechol O-monosulfate products that were identified by reversed phase HPLC and by LC-MS/MS. The various SULT isoforms produced the two isomers in different proportions. Two-dimensional (1)H and (13)C NMR assigned the two regioisomers of benzo[a]pyrene-7,8-catechol monosulfate as 8-hydroxy-benzo[a]pyrene-7-O-sulfate (M1) and 7-hydroxy-benzo[a]pyrene-8-O-sulfate (M2), respectively. The kinetic profiles of three SULTs were different. SULT1A1 gave the highest catalytic efficiency (k(cat)/K(m)) and yielded a single isomeric product corresponding to M1. By contrast, SULT1E1 showed distinct substrate inhibition and formed both M1 and M2. Based on expression levels, catalytic efficiency, and the fact that the lung cells only produce M1, it is concluded that the major isoform that can intercept benzo[a]pyrene-7,8-catechol is SULT1A1.
Highlights
Benzo[a]pyrene-7,8-dione is a genotoxic metabolite produced by aldo-keto reductases
Among the six different SULT isoforms, SULT1A1, -1A3, and -1E1 were expressed at the mRNA level (Fig. 2). This screen showed that SULT1A3 was the most abundant at the mRNA level in human lung cells followed by SULT1A1
We have shown that Polycyclic aromatic hydrocarbons (PAH) o-quinones are enzymatically redox-cycled at a robust rate by recombinant NQO1 and by aldo-keto reductases (AKR) themselves and that human catechol-O-methyl transferase (COMT) can O-methylate a series of PAH-catechols [21, 26]
Summary
Benzo[a]pyrene-7,8-dione is a genotoxic metabolite produced by aldo-keto reductases. Results: SULT1A1 is identified as the major SULT in lung cells involved in the detoxication of the corresponding catechol. Catalytic efficiency, and the fact that the lung cells only produce M1, it is concluded that the major isoform that can intercept benzo[a]pyrene-7,8-catechol is SULT1A1. The third pathway is termed the o-quinone pathway catalyzed by aldo-keto reductases (AKR) In this pathway, B[a]P-7,8-trans-dihydrodiol is oxidized by AKR1A1 and AKR1C1–1C4 to yield a ketol that spontaneously rearranges to form B[a]P-7,8-catechol, which is not stable and undergoes autooxidation to yield benzo[a]pyrene7,8-dione (B[a]P-7,8-dione) and the generation of reactive oxygen species (ROS) [13,14,15] (Fig. 1). In human lung adenocarcinoma (A549) cells with high constitutive expression of AKRs, the metabolic activation of B[a]P-7,8-trans-dihydrodiol to B[a]P7,8-dione by AKRs led to the formation ROS and 8-oxo-dGuo adducts in cellular DNA, which were exacerbated by the presence of a catechol-O-methyl transferase (COMT) inhibitor [25]. It is imperative to investigate whether sulfonation of B[a]P-7,8-catechol by SULTs is a detoxication pathway for B[a]P-7,8-dione that will limit its ability to redox cycle
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