Glycosylation of polycyclic aromatic hydrocarbons by UDP‐glycosyltransferase 3A2 (UGT3A2) and aerodigestive tract tissues

Abstract

Smoking is a primary risk factor for the development of lung and tobacco‐related cancers. There are over 70 carcinogens in tobacco smoke, and they belong to multiple chemical classes including polycyclic aromatic hydrocarbons (PAHs). The UDP‐glycosyltransferase (UGT) family of enzymes play a key role in the detoxification of several major tobacco carcinogens by glucuronidation, using UDP‐glucuronic acid as cosubstrate. While the UGT 1As, 2As, and 2Bs are well known to detoxify simple and complex PAHs, the UGT3A subfamily has been understudied in the detoxification of PAHs. UGT3A2 uses UDP‐glucose and UDP‐xylose as cosubstrates and is active against 1‐hydroxypyrene, a biomarker for PAH exposure. Our previous research has shown that UGT3A2 is expressed in the aerodigestive tract and is active against simple hydroxylated PAHs. The purpose of the present study was to determine the kinetic parameters of UGT3A2 against complex PAHs and determine glycosylation in aerodigestive tract tissues using UDP‐glucose and UDP‐xylose. Microsomes prepared from UGT3A2 overexpressing HEK293 cells exhibited glycosylation activity against complex PAHs including benzo[a]pyrene (B[a]P)‐7,8‐diol, B[a]P‐9,10‐diol, DB[a, l]P‐11,12‐diol, and 5‐methylchrysene‐1,2‐diol, with Km's ranging from 96 to 1250 μM using UDP‐glucose. The Km's were significantly lower when using UDP‐xylose, with Km's ranging from 120 to 168 μM for the same substrates. The Vmax/Km ratio of UDP‐xylose/UDP‐glucose ranged from 0.71 to 4.0. To determine the physiological significance and relevance of UDP‐xylose‐ and UDP‐glucose‐dependent glycosylation, activity assays were performed in tissues where UGT3A2 is expressed using 1‐hydroxypyrene as substrate. Using UDP‐glucose as the cosubstrate, glycosylation rates of 3.5, 5.2, and 13 pmol·min−1·mg−1 were observed for liver, intestine and kidney microsomes, respectively. The rates were higher using UDP‐xylose, with rates of 22, 9.3, and 20 pmol·min−1·mg−1 for liver, intestine and kidney, respectively. The glycosylation rate ratio for UDP‐xylose/UDP‐glucose ranged from 1.6 to 6.3 in the three tissues, with liver exhibiting the highest ratio. In addition, detectable glycosylation activity was observed in various aerodigestive tract tissues including esophagus, tonsil, bronchus, floor of mouth, tongue, trachea, and larynx, with tonsil exhibiting the highest relative activity with both UDP‐sugars. Incubations with UDP‐xylose resulted in higher levels of glycosylation activity than that observed with UDP‐glucose in esophagus, tonsil, floor of mouth, and larynx. These data suggest that UGT3A2 is playing an important role in the detoxification of PAHs in tobacco target tissues, with UDP‐xylose being the preferred cosubstrate in most tissues.Support or Funding InformationThis study is supported by the National Institute of Environmental Health Sciences (NIEHS) of the National Institutes of Health to P Lazarus (Grant R01‐ES025460), a NIEHS Research Supplement to Promote Diversity in Health‐Related Research to AG Vergara (Grant R01‐ES025460‐02S1), and a grant from the Health Sciences and Services Authority of Spokane, WA (Grant WSU002292).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.