초록
<P>Glutathione-dependent enzymes play important protective, repair, or metabolic roles in cells. In particular, enzymes in the glutathione <I>S</I>-transferase (GST) superfamily function in stress responses, defense systems, or xenobiotic detoxification. Here, we identify novel features of bacterial GSTs that cleave β-aryl ether bonds typically found in plant lignin. Our data reveal several original features of the reaction cycle of these GSTs, including stereospecific substrate recognition and stereoselective formation of β-<I>S</I>-thioether linkages. Products of recombinant GSTs (LigE, LigP, and LigF) are β-<I>S</I>-glutathionyl-α-keto-thioethers that are degraded by a β-<I>S</I>-thioetherase (LigG). All three Lig GSTs produced the ketone product (β-<I>S</I>-glutathionyl-α-veratrylethanone) from an achiral side chain-truncated model substrate (β-guaiacyl-α-veratrylethanone). However, when β-etherase assays were conducted with a racemic model substrate, β-guaiacyl-α-veratrylglycerone, LigE- or LigP-catalyzed reactions yielded only one of two potential product (β-<I>S</I>-glutathionyl-α-veratrylglycerone) epimers, whereas the other diastereomer (differing in configuration at the β-position (<I>i.e.</I> its β-epimer)) was produced only in the LigF-catalyzed reaction. Thus, β-etherase catalysis causes stereochemical inversion of the chiral center, converting a β(<I>R</I>)-substrate to a β(<I>S</I>)-product (LigE and LigP), and a β(<I>S</I>)-substrate to a β(<I>R</I>)-product (LigF). Further, LigG catalyzed glutathione-dependent β-<I>S</I>-thioether cleavage with β-<I>S</I>-glutathionyl-α-veratrylethanone and with β(<I>R</I>)-configured β-<I>S</I>-glutathionyl-α-veratrylglycerone but exhibited no or significantly reduced β-<I>S</I>-thioether-cleaving activity with the β(<I>S</I>)-epimer, demonstrating that LigG is a stereospecific β-thioetherase. We therefore propose that multiple Lig enzymes are needed in this β-aryl etherase pathway in order to cleave the racemic β-ether linkages that are present in the backbone of the lignin polymer.</P>