Roles of two glutathione S-transferases in the final step of the β-aryl ether cleavage pathway in Sphingobium sp. strain SYK-6

Abstract

ABSTRACTSphingobium sp. strain SYK-6 is an alphaproteobacterial degrader of lignin-derived aromatic compounds, which can degrade all the stereoisomers of β-aryl ether-type compounds. SYK-6 cells convert four stereoisomers of guaiacylglycerol-β-guaiacyl ether (GGE) into two enantiomers of α-(2-methoxyphenoxy)-β-hydroxypropiovanillone (MPHPV) through GGE α-carbon atom oxidation by stereoselective Cα-dehydrogenases encoded by ligD, ligL, and ligN. The ether linkages of the resulting MPHPV enantiomers are cleaved by stereoselective glutathione S-transferases (GSTs) encoded by ligF, ligE, and ligP, generating (βRβS)-α-glutathionyl-β-hydroxypropiovanillone (GS-HPV) and guaiacol. To date, it has been shown that the gene products of ligG and SLG_04120 (ligQ), both encoding GST, catalyze glutathione removal from (βRβS)-GS-HPV, forming achiral β-hydroxypropiovanillone. In this study, we characterized the enzyme properties of LigG and LigQ and elucidated their roles in β-aryl ether catabolism. Purified LigG showed an approximately 300-fold higher specific activity for (βR)-GS-HPV than that for (βS)-GS-HPV, whereas purified LigQ showed an approximately six-fold higher specific activity for (βS)-GS-HPV than that for (βR)-GS-HPV. Analyses of mutants of ligG, ligQ, and both genes revealed that SYK-6 converted (βR)-GS-HPV using both LigG and LigQ, whereas only LigQ was involved in converting (βS)-GS-HPV. Furthermore, the disruption of both ligG and ligQ was observed to lead to the loss of the capability of SYK-6 to convert MPHPV. This suggests that GSH removal from GS-HPV catalyzed by LigG and LigQ, is essential for cellular GSH recycling during β-aryl ether catabolism.IMPORTANCEThe β-aryl ether linkage is most abundant in lignin, comprising 45%–62% of all intermonomer linkages in lignin; thus, cleavage of the β-aryl ether linkage together with the subsequent degradation process is considered the essential step in lignin biodegradation. The enzyme genes for β-aryl ether cleavage are useful for decomposing high-molecular-weight lignin, converting lignin-derived aromatic compounds into value-added products, and modifying lignin structures in plants to reduce lignin recalcitrance. In this study, we uncovered the roles of the two glutathione S-transferase genes, ligG and ligQ, in the conversion of GS-HPV isomers, which are generated in the β-aryl ether cleavage pathway in SYK-6. Adding our current results to previous findings allowed us to have a whole picture of the β-aryl ether cleavage system in SYK-6.