Methanethiol (MT) is a sulfur-containing compound produced during dimethylsulfoniopropionate (DMSP) degradation by marine bacteria. The C-S bond of MT can be cleaved by methanethiol oxidases (MTOs) to release a sulfur atom. However, the cleaving process remains unclear, and the species of sulfur product is uncertain. It has long been assumed that MTOs produce hydrogen sulfide (H2S) from MT. Herein, we studied the MTOs in the Rhodobacteraceae family-whose members are important DMSP degraders ubiquitous in marine environments. We identified 57 MTOs from 1,904 Rhodobacteraceae genomes. These MTOs were grouped into two major clusters. Cluster 1 members share three conserved cysteine residues, while cluster 2 members contain one conserved cysteine residue. We examined the products of three representative MTOs both in vitro and in vivo. All of them produced sulfane sulfur other than H2S from MT. Their conserved cysteines are substrate-binding sites in which the MTO-S-S-CH3 complex is formed. This finding clarified the sulfur product of MTOs and enlightened the MTO-catalyzing process. Moreover, this study connected DMSP degradation with sulfane sulfur metabolism, filling a critical gap in the DMSP degradation pathway and representing new knowledge in the marine sulfur cycle field.
OBJECTIVE: This study overthrows a long-time assumption that methanethiol oxidases (MTOs) cleave the C-S bond of methanethiol to produce both H2S and H2O2-the former is a strong reductant and the latter is a strong oxidant. From a chemistry viewpoint, this reaction is difficult to happen. Investigations on three representative MTOs indicated that sulfane sulfur (S0) was the direct product, and no H2O2 was produced. Finally, the products of MTOs were corrected to be S0 and H2O. This finding connected dimethylsulfoniopropionate (DMSP) degradation with sulfane sulfur metabolism, filling a critical gap in the DMSP degradation pathway and representing new knowledge in the marine sulfur cycle field.

Methanethiol is a toxic gas produced through bacterial degradation of sulfur-containing amino acids. Applying a novel enzymatic assay, we here identified a methanethiol oxidase (MTO) that catalyzes the degradation of methanethiol in the nematode Caenorhabditis elegans (C. elegans). The corresponding protein, Y37A1B.5, previously characterized as a C. elegans ortholog of human selenium-binding protein 1 (SELENBP1), was renamed SEMO-1 (SELENBP1 ortholog with methanethiol oxidase activity). Worms rendered deficient in SEMO-1 not only showed decreased hydrogen sulfide production from methanethiol catabolism but they were also more resistant to oxidative stress and had an elevated life span. In contrast, resistance to selenite was significantly lowered in SEMO-1-deficient worms. Naturally occurring mutations of human SELENBP1 were introduced to recombinant SEMO-1 through site-directed mutagenesis and resulted in loss of its MTO activity, indicating a similar enzymatic mechanism for SELENBP1 and SEMO-1. In summary, SEMO-1 confers resistance to toxic selenite and the ability to metabolize toxic methanethiol. These beneficial effects might be a trade-off for its negative impact on C. elegans life span.

Methanethiol, a gas with the characteristic smell of rotten cabbage, is a product of microbial methionine degradation. In the human body, methanethiol originates primarily from bacteria residing in the lumen of the large intestine. Selenium-binding protein 1 (SELENBP1), a marker protein of mature enterocytes, has recently been identified as a methanethiol oxidase (MTO). It catalyzes the conversion of methanethiol to hydrogen sulfide (H2S), hydrogen peroxide (H2O2) and formaldehyde. Here, human Caco-2 intestinal epithelial cells were subjected to enterocyte-like differentiation, followed by analysis of SELENBP1 levels and MTO activity. To that end, we established a novel coupled assay to assess MTO activity mimicking the proximity of microbiome and intestinal epithelial cells in vivo. The assay is based on in situ-generation of methanethiol as catalyzed by a bacterial recombinant l-methionine gamma-lyase (MGL), followed by detection of H2S and H2O2. Applying this assay, we verified the loss and impairment of MTO function in SELENBP1 variants (His329Tyr; Gly225Trp) previously identified in individuals with familial extraoral halitosis. MTO activity was strongly enhanced in Caco-2 cells upon enterocyte differentiation, in parallel with increased SELENBP1 levels. This suggests that mature enterocytes located at the tip of colonic crypts are capable of eliminating microbiome-derived methanethiol.