Journal Article

Role of arginine 216 in catalytic activity of murine Alpha class glutathione transferases mGSTAl-1 and mGSTA2-2 toward carcinogenic diol epoxides of polycyclic aromatic hydrocarbons

Ajai Pal, Yijun Gu, Christian Herzog, Sanjay K. Srivastava, Piotr Zimniak, Xinhua Ji and Shivendra V. Singh

in Carcinogenesis

Volume 22, issue 8, pages 1301-1305
Published in print August 2001 | ISSN: 0143-3334
Published online August 2001 | e-ISSN: 1460-2180 | DOI: https://dx.doi.org/10.1093/carcin/22.8.1301
Role of arginine 216 in catalytic activity of murine Alpha class glutathione transferases mGSTAl-1 and mGSTA2-2 toward carcinogenic diol epoxides of polycyclic aromatic hydrocarbons

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Murine class Alpha glutathione (GSH) transferase A1-1 (mGSTA1-1) is unique among mammalian Alpha class GSTs due to its exceptionally high catalytic activity toward (+)-anti-7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene [(+)-anti-BPDE], which is the activated metabolite of an environmentally relevant carcinogen, benzo[a] pyrene (BP). However, the molecular basis for high catalytic activity of mGSTA1-1 toward (+)-anti-BPDE is not clear. In the present study, we demonstrate that an arginine residue at position 216, which is conserved in some but not all mammalian class Alpha GSTs, plays an important role in catalytic activity of mGSTA1-1 toward (+)-anti-BPDE and carcinogenic diol epoxides of other environmentally relevant polycyclic aromatic hydrocarbons (PAHs). The catalytic efficiency (kcat/Km) of mGSTA1-1 for the GSH conjugation of (+)-anti-BPDE (108/mM/s) was reduced by about 58% upon replacement of arginine 216 with alanine (R216A). This was mainly due to a significantly lower Vmax for the R216A mutant of mGSTA1-1 compared with wild-type mGSTA1-1. The R216A mutation also resulted in a statistically significant reduction (>70%) in specific activity of mGSTA1-1 toward racemic anti-diol epoxides of chrysene and benzo[c]phenanthrene (anti-CDE and anti-B[c]PDE, respectively). The catalytic activity of mGSTA2-2, which is a close structural homologue of mGSTA1-1, was also reduced upon R216A mutation. The results of the present study clearly indicate that an arginine residue at position 216 is critical for catalytic activity of mGSTA1-1 and mGSTA2-2 toward carcinogenic diol epoxide metabolites of various PAHs that are abundant in the environment and suspected human carcinogens.

Keywords: (+)-anti-BPDE, (+)-anti-7,8-dihydroxy-9,10-epoxy-7,8,9, 10-tetrahydrobenzo[a]pyrene; anti-B[c]PDE, anti-3,4-dihydroxy-1,2-epoxy-1,2,3,4-tetrahydrobenzo[c]phenanthrene; anti-CDE, anti-1,2-dihydroxy-3,4-epoxy-1,2,3,4-tetrahydrochrysene; BP, benzo[a]pyrene; GSBpd, GSH conjugate of (+)-anti-BPDE; GSH, glutathione; GST, glutathione transferase; mGSTA1-1, murine class Alpha GST isoenzyme A1-1; mGSTA1-1/R216A; arginine 216→alanine mutant of mGSTA1-1; mGSTA1-1–GSH, mGSTA1-1 in complex with GSH; mGSTA1-1–GSBpd, mGSTA1-1 in complex with GSBpd; mGSTA2-2, murine class Alpha GST isoenzyme A2-2; mGSTA2-2/R216A, arginine 216→alanine mutant of mGSTA2-2; hGSTA1-1, human class Alpha GST isoenzyme A1-1; H-site, hydrophobic substrate binding site; PAHs, polycyclic aromatic hydrocarbons; R216A, enzyme with mutation of arginine 216 with alanine.

Journal Article.  4436 words.  Illustrated.

Subjects: Clinical Cytogenetics and Molecular Genetics

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