The DinB Superfamily Includes Novel Mycothiol, Bacillithiol, and Glutathione S-Transferases

Abstract

The superfamily of glutathione S-transferases has been the subject of extensive study but Actinobacteria produce mycothiol (MSH) in place of glutathione and no mycothiol S-transferase (MST) has been identified. Using mycothiol and monochlorobimane as substrates a MST activity was detected in extracts of Mycobacterium smegmatis and purified sufficiently to allow identification of MSMEG_0887, a member the DUF664 family of the DinB superfamily, as the MST. The identity of the M. smegmatis and homologous Mycobacterium tuberculosis (Rv0443) enzymes was confirmed by cloning and the expressed proteins were found to be active with MSH but not bacillithiol (BSH) or glutathione (GSH). Bacillus subtilis YfiT is another member of the DinB superfamily but this bacterium produces BSH. The YfiT protein was shown to have S-transferase activity with monochlorobimane when assayed with BSH but not with MSH or GSH. Enterococcus faecalis EF_3021 shares some homology with MSMEG_0887 but this organism produces GSH but not MSH or BSH. Cloned and expressed EF_0321 was active with monochlorobimane and GSH but not with MSH or BSH. MDMPI_2 is another member of the DinB superfamily and has been previously shown to have mycothiol-dependent maleylpyruvate isomerase activity. Three of the eight families of the DinB superfamily include proteins shown to catalyze thiol-dependent metabolic or detoxification activities. Since more than two-thirds of the sequences assigned to the DinB superfamily are members of these families it seems likely that such activity is dominant in the DinB superfamily.

Detoxification of toxins by bacillithiol in Staphylococcus aureus

“…When we followed the fate of the fluorescent alkylating agent monochlorobimane, which is less chemically reactive and thus would require a BST, analogous to a GST, to catalyse its conjugation to BSH, the mercapturic acid of the bimane was detected intracellularly and extracellularly, indicating the presence of a BST, which catalyses the conjugation of monochlorobimane with BSH to produce BSmB. Recently, we have identified and characterized a B. subtilis BST which is able to catalyse the reaction of monochlorobimane with BSH; whether the S. aureus homologue does the same remains to be seen (Newton et al, 2011).…”

Comparative Proteomic Analysis of Streptomyces lividans Wild-Type andppkMutant Strains Reveals the Importance of Storage Lipids for Antibiotic Biosynthesis

“…A protein (SCO7269) belonging to the DinB (DNA damage-inducible) superfamily (44) and bearing a mycothiol-dependent maleylpyruvate isomerase Nterminal domain was found to be present in larger amounts in the ppk mutant than in the wt strain. Proteins of this family were shown to catalyze thiol-dependent detoxification reactions (45).…”

Novel l -Cysteine-Dependent Maleylpyruvate Isomerase in the Gentisate Pathway of Paenibacillus sp. Strain NyZ101

“…During the preparation of the present study the characterization of three additional enzymes in DinB superfamily was reported to suggest that thiol-dependent S-transferase activity is dominant in this family, including MSH, bacillithiol (BSH), and GSH S-transferases (32). In particular, B. subtilis YfiT, mentioned above, was shown to have BSH S-transferase activity (32). As shown in the phylogenetic tree of the DinB superfamily in Fig.…”

“…The complete loss of enzyme activity in three BagL mutants indicated that the three histidines are necessary for its activity and probably function in binding with Ni 2ϩ . During the preparation of the present study the characterization of three additional enzymes in DinB superfamily was reported to suggest that thiol-dependent S-transferase activity is dominant in this family, including MSH, bacillithiol (BSH), and GSH S-transferases (32). In particular, B. subtilis YfiT, mentioned above, was shown to have BSH S-transferase activity (32).…”