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The factors shaping host-parasite interactions and epibiont communities in the variable rocky intertidal zone are poorly understood. California mussels, Mytilus californianus, are colonized by endolithic cyanobacterial parasites which erode the host shell. These cyanobacteria become mutualistic under certain abiotic conditions because shell erosion can protect mussels from thermal stress. How parasitic shell erosion affects or is affected by epibiotic microbial communities on mussel shells and the context dependency of these interactions is unknown. We used transplant experiments to characterize assemblages of epibiotic bacteria and endolithic parasites on mussel shells across intertidal elevation gradients. We hypothesized that living mussels, and associated epibacterial communities, could limit colonization and erosion by endolithic cyanobacteria compared to empty mussel shells. We hypothesized that shell erosion would be associated with compositional shifts in the epibacterial community and tidal elevation. We found that living mussels experienced less shell erosion than empty shells, demonstrating potential biotic regulation of endolithic parasites. Increased shell erosion was not associated with a distinct epibacterial community and was decoupled from the relative abundance of putatively endolithic taxa. Our findings suggest that epibacterial community structure is not directly impacted by the dynamic symbiosis between endolithic cyanobacteria and mussels throughout the rocky intertidal zone.
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Selenium is an essential micronutrient, but its presence in biology has been limited to protein and nucleic acid biopolymers. The recent identification of a biosynthetic pathway for selenium-containing small molecules suggests that there is a larger family of selenometabolites that remains to be discovered. Here we identify a recently evolved branch of abundant and uncharacterized metalloenzymes that we predict are involved in selenometabolite biosynthesis using a bioinformatic search strategy that relies on the mapping of composite active site motifs. Biochemical studies confirm this prediction and show that these enzymes form an unusual C-Se bond onto histidine, thus giving rise to a distinct selenometabolite and potent antioxidant that we have termed ovoselenol. Aside from providing insights into the evolution of this enzyme class and the structural basis of C-Se bond formation, our work offers a blueprint for charting the microbial selenometabolome in the future.
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Keratinicyclins and keratinimicins are recently discovered glycopeptide antibiotics. Keratinimicins show broad-spectrum activity against Gram-positive bacteria, while keratinicyclins form a new chemotype by virtue of an unusual oxazolidinone moiety and exhibit specific antibiosis against Clostridioides difficile. Here we report the mechanism of action of keratinicyclin B (KCB). We find that steric constraints preclude KCB from binding peptidoglycan termini. Instead, KCB inhibits C. difficile growth by binding wall teichoic acids (WTAs) and interfering with cell wall remodeling. A computational model, guided by biochemical studies, provides an image of the interaction of KCB with C. difficile WTAs and shows that the same H-bonding framework used by glycopeptide antibiotics to bind peptidoglycan termini is used by KCB for interacting with WTAs. Analysis of KCB in combination with vancomycin (VAN) shows highly synergistic and specific antimicrobial activity, and that nanomolar combinations of the two drugs are sufficient for complete growth inhibition of C. difficile, while leaving common commensal strains unaffected.
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Antibacterianos , Clostridioides difficile , Pruebas de Sensibilidad Microbiana , Clostridioides difficile/efectos de los fármacos , Antibacterianos/farmacología , Antibacterianos/química , Vancomicina/farmacología , Vancomicina/química , Pared Celular/efectos de los fármacos , Pared Celular/metabolismo , Ácidos Teicoicos/metabolismo , Peptidoglicano/metabolismo , Peptidoglicano/química , Quimioterapia Combinada , Péptidos Cíclicos , LipopéptidosRESUMEN
Selenium is an essential micronutrient, but its presence in biology has been limited to protein and nucleic acid biopolymers. The recent identification of the first biosynthetic pathway for selenium-containing small molecules suggests that there is a larger family of selenometabolites that remains to be discovered. Using a bioinformatic search strategy that relies on mapping of composite active site motifs, we identify a recently evolved branch of abundant and uncharacterized metalloenzymes that we predict are involved in selenometabolite biosynthesis. Biochemical studies confirm this prediction and show that these enzymes form an unusual C-Se bond onto histidine, thus giving rise to a novel selenometabolite and potent antioxidant that we have termed ovoselenol. Aside from providing insights into the evolution of this enzyme class and the structural basis of C-Se bond formation, our work offers a blueprint for charting the microbial selenometabolome in the future.
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Incarcerated pregnant people face significant barriers when seeking health care services in prisons and jails, but little is known about their transitions from state prison health care systems to outside hospitals. This project analyzed current policies and procedures for care transitions for incarcerated people and presents policy recommendations to address issues of concern. We conducted in-depth interviews with stakeholders at a state prison, academic hospital, and private hospital to identify the barriers and facilitators to care transitions. Themes emerging from these interviews were operational, including medical records, communication, and education; and structural, including implicit biases and care of marginalized groups. These findings are likely applicable to similar facilities throughout the United States. A multipronged, interdisciplinary approach is needed to address challenges of care transitions.
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Prisioneros , Femenino , Embarazo , Humanos , Estados Unidos , Evaluación de Necesidades , Transferencia de Pacientes , Prisiones , Atención a la SaludRESUMEN
Selenium (Se) is an essential micronutrient that is found naturally in proteins, nucleic acids, and natural products. Unlike selenoproteins and selenonucleic acids, little is known about the structures of biosynthetic enzymes that incorporate Se into small molecules. Here, we report the X-ray crystal structure of SenB, the first known Se-glycosyltransferase that was recently found to be involved in the biosynthesis of the Se-containing metabolite selenoneine. SenB catalyzes C-Se bond formation using selenophosphate and an activated uridine diphosphate sugar as a Se and glycosyl donor, respectively, making it the first known selenosugar synthase and one of only four bona fide C-Se bond-forming enzymes discovered to date. Our crystal structure, determined to 2.25 Å resolution, reveals that SenB is a type B glycosyltransferase, displaying the prototypical fold with two globular Rossmann-like domains and a catalytic interdomain cleft. By employing complementary structural biology techniques, we find that SenB undergoes both local and global substrate-induced conformational changes, demonstrating a significant increase in α-helicity and a transition to a more compact conformation. Our results provide the first structure of SenB and set the stage for further biochemical characterization in the future.
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Selenio , Selenio/metabolismo , Glicosiltransferasas , Ligandos , Selenoproteínas , Cristalografía por Rayos XRESUMEN
Nitration reactions are crucial for many industrial syntheses; however, current protocols lack site specificity and employ hazardous chemicals. The noncanonical cytochrome P450 enzymes RufO and TxtE catalyze the only known direct aromatic nitration reactions in nature, making them attractive model systems for the development of analogous biocatalytic and/or biomimetic reactions that proceed under mild conditions. While the associated mechanism has been well-characterized in TxtE, much less is known about RufO. Herein we present the first structure of RufO alongside a series of computational and biochemical studies investigating its unusual reactivity. We demonstrate that free l-tyrosine is not readily accepted as a substrate despite previous reports to the contrary. Instead, we propose that RufO natively modifies l-tyrosine tethered to the peptidyl carrier protein of a nonribosomal peptide synthetase encoded by the same biosynthetic gene cluster and present both docking and molecular dynamics simulations consistent with this hypothesis. Our results expand the scope of direct enzymatic nitration reactions and provide the first evidence for such a modification of a peptide synthetase-bound substrate. Both of these insights may aid in the downstream development of biocatalytic approaches to synthesize rufomycin analogues and related drug candidates.
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Sistema Enzimático del Citocromo P-450 , Nitratos , Nitratos/metabolismo , Sistema Enzimático del Citocromo P-450/metabolismo , Simulación de Dinámica Molecular , Tirosina , Especificidad por SustratoRESUMEN
The emergence of multidrug-resistant pathogens poses a threat to public health and requires new antimicrobial agents. As the archetypal glycopeptide antibiotic (GPA) used against drug-resistant Gram-positive pathogens, vancomycin provides a promising starting point. Peripheral alterations to the vancomycin scaffold have enabled the development of new GPAs. However, modifying the core remains challenging due to the size and complexity of this compound family. The recent successful chemoenzymatic synthesis of vancomycin suggests that such an approach can be broadly applied. Herein, we describe the expansion of chemoenzymatic strategies to encompass type II GPAs bearing all aromatic amino acids through the production of the aglycone analogue of keratinimicin A, a GPA that is 5-fold more potent than vancomycin against Clostridioides difficile. In the course of these studies, we found that the cytochrome P450 enzyme OxyBker boasts both broad substrate tolerance and remarkable selectivity in the formation of the first aryl ether cross-link on the linear peptide precursors. The X-ray crystal structure of OxyBker, determined to 2.8 Å, points to structural features that may contribute to these properties. Our results set the stage for using OxyBker broadly as a biocatalyst toward the chemoenzymatic synthesis of diverse GPA analogues.
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Antibacterianos , Vancomicina , Vancomicina/química , Antibacterianos/química , Glicopéptidos/química , Sistema Enzimático del Citocromo P-450/metabolismo , PéptidosRESUMEN
Kelp are important primary producers that are colonized by diverse microbes that can have both positive and negative effects on their hosts. The kelp microbiome could support the burgeoning kelp cultivation sector by improving host growth, stress tolerance, and resistance to disease. Fundamental questions about the cultivated kelp microbiome still need to be addressed before microbiome-based approaches can be developed. A critical knowledge gap is how cultivated kelp microbiomes change as hosts grow, particularly following outplanting to sites that vary in abiotic conditions and microbial source pools. In this study we assessed if microbes that colonize kelp in the nursery stage persist after outplanting. We characterized microbiome succession over time on two species of kelp, Alaria marginata and Saccharina latissima, outplanted to open ocean cultivation sites in multiple geographic locations. We tested for host-species specificity of the microbiome and the effect of different abiotic conditions and microbial source pools on kelp microbiome stability during the cultivation process. We found the microbiome of kelp in the nursery is distinct from that of outplanted kelp. Few bacteria persisted on kelp following outplanting. Instead, we identified significant microbiome differences correlated with host species and microbial source pools at each cultivation site. Microbiome variation related to sampling month also indicates that seasonality in host and/or abiotic factors may influence temporal succession and microbiome turnover in cultivated kelps. This study provides a baseline understanding of microbiome dynamics during kelp cultivation and highlights research needs for applying microbiome manipulation to kelp cultivation.
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Kelp , Microbiota , Phaeophyceae , BacteriasRESUMEN
Enzyme reactivity is often enhanced by changes in oxidation state, spin state, and metal-ligand covalency of associated metallocofactors. The development of spectroscopic methods for studying these processes coincidentally with structural rearrangements is essential for elucidating metalloenzyme mechanisms. Herein, we demonstrate the feasibility of collecting X-ray emission spectra of metalloenzyme crystals at a third-generation synchrotron source. In particular, we report the development of a von Hamos spectrometer for the collection of Fe Kß emission optimized for analysis of dilute biological samples. We further showcase its application in crystals of the immunosuppressive heme-dependent enzyme indoleamine 2,3-dioxygenase. Spectra from protein crystals in different states were compared with relevant reference compounds. Complementary density functional calculations assessing covalency support our spectroscopic analysis and identify active site conformations that correlate to high- and low-spin states. These experiments validate the suitability of an X-ray emission approach for determining spin states of previously uncharacterized metalloenzyme reaction intermediates.
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Hemo , Metaloproteínas , Hemo/metabolismo , Espectrometría por Rayos X , Metales , Dominio CatalíticoRESUMEN
Dietary fibers are potent modulators of immune responses that can restrain inflammation in multiple disease contexts. However, dietary fibers encompass a biochemically diverse family of carbohydrates, and it remains unknown how individual fiber sources influence immunity. In a direct comparison of four different high-fiber diets, we demonstrate a potent ability of guar gum to delay disease and neuroinflammation in experimental autoimmune encephalomyelitis, a T cell-mediated mouse model of multiple sclerosis. Guar gum-specific alterations to the microbiota are limited, and disease protection appears to be independent of fiber-induced increases in short-chain fatty acid levels or regulatory CD4+ T cells. Instead, CD4+ T cells of guar gum-supplemented mice are less encephalitogenic due to reduced activation, proliferation, Th1 differentiation, and altered migratory potential. These findings reveal specificity in the host response to fiber sources and define a pathway of fiber-induced immunomodulation that protects against pathologic neuroinflammation.
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Cyamopsis , Encefalomielitis Autoinmune Experimental , Animales , Cyamopsis/metabolismo , Dieta , Fibras de la Dieta/farmacología , Fibras de la Dieta/uso terapéutico , Encefalomielitis Autoinmune Experimental/tratamiento farmacológico , Galactanos , Mananos , Ratones , Gomas de PlantasRESUMEN
Hydroxyalkylquinolines (HAQs) are ubiquitious natural products but their interactions with associated protein targets remain elusive. We report X-ray crystal structures of two HAQs in complex with dihydroorotate dehydrogenase (DHODH). Our results reveal the structural basis of DHODH inhibition by HAQs and open the door to downstream structure-activity relationship studies.
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The cyanobacterial enzyme CylK assembles the cylindrocyclophane natural products by performing two unusual alkylation reactions, forming new carbon-carbon bonds between aromatic rings and secondary alkyl halide substrates. This transformation is unprecedented in biology, and the structure and mechanism of CylK are unknown. Here, we report X-ray crystal structures of CylK, revealing a distinctive fusion of a Ca2+-binding domain and a ß-propeller fold. We use a mutagenic screening approach to locate CylK's active site at its domain interface, identifying two residues, Arg105 and Tyr473, that are required for catalysis. Anomalous diffraction datasets collected with bound bromide ions, a product analog, suggest that these residues interact with the alkyl halide electrophile. Additional mutagenesis and molecular dynamics simulations implicate Asp440 in activating the nucleophilic aromatic ring. Bioinformatic analysis of CylK homologs from other cyanobacteria establishes that they conserve these key catalytic amino acids, but they are likely associated with divergent reactivity and altered secondary metabolism. By gaining a molecular understanding of this unusual biosynthetic transformation, this work fills a gap in our understanding of how alkyl halides are activated and used by enzymes as biosynthetic intermediates, informing enzyme engineering, catalyst design, and natural product discovery.
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Cianobacterias , Alquilación , Carbono/metabolismo , Catálisis , Dominio Catalítico , Cristalografía por Rayos X , Cianobacterias/genética , Cianobacterias/metabolismoRESUMEN
Enzymes are biological catalysts whose dynamics enable their reactivity. Visualizing conformational changes, in particular, is technically challenging, and little is known about these crucial atomic motions. This is especially problematic for understanding the functional diversity associated with the radical S-adenosyl-L-methionine (SAM) superfamily whose members share a common radical mechanism but ultimately catalyze a broad range of challenging reactions. Computational chemistry approaches provide a readily accessible alternative to exploring the time-resolved behavior of these enzymes that is not limited by experimental logistics. Here, we review the application of molecular docking, molecular dynamics, and density functional theory, as well as hybrid quantum mechanics/molecular mechanics methods to the study of these enzymes, with a focus on understanding the mechanistic dynamics associated with turnover.
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Modelos Moleculares , S-Adenosilmetionina/química , Acetiltransferasas , Teoría Funcional de la Densidad , Conformación Molecular , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Estructura Molecular , Unión Proteica , Teoría Cuántica , Vitamina B 12/químicaRESUMEN
It is unclear how host-associated microbial communities will be affected by future environmental change. Characterizing how microbiota differ across sites with varying environmental conditions and assessing the stability of the microbiota in response to abiotic variation are critical steps towards predicting outcomes of environmental change. Intertidal organisms are valuable study systems because they experience extreme variation in environmental conditions on tractable timescales such as tide cycles and across small spatial gradients in the intertidal zone. Here we show a widespread intertidal macroalgae, Fucus distichus, hosts site-specific microbiota over small (meters to kilometres) spatial scales. We demonstrate stability of site-specific microbial associations by manipulating the host environment and microbial species pool with common garden and reciprocal transplant experiments. We hypothesized that F. distichus microbiota would readily shift to reflect the contemporary environment due to selective filtering by abiotic conditions and/or colonization by microbes from the new environment or nearby hosts. Instead, F. distichus microbiota was stable for days after transplantation in both the laboratory and field. Our findings expand the current understanding of microbiota dynamics on an intertidal foundation species. These results may also point to adaptations for withstanding short-term environmental variation, in hosts and/or microbes, facilitating stable host-microbial associations.
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Fucus , Microbiota , Algas Marinas , Adaptación FisiológicaRESUMEN
Seaweed-associated microbiota are essential for the health and resilience of nearshore ecosystems, marine biogeochemical cycling, and host health. Yet much remains unknown about the ecology of seaweed-microbe symbioses. In this study, we quantified fine-scale patterns of microbial community structure across distinct anatomical regions of the kelp Laminaria setchellii. These anatomical regions represent a gradient of tissue ages: perennial holdfasts can be several years old, whereas stipe epicortex and blades are younger annual structures. Within blades, new growth occurs at the base, while the blade tips may be several months old and undergoing senescence. We hypothesized that microbial communities will differ across anatomical regions (holdfast, stipe, blade base, and blade tip), such that younger tissues will harbor fewer microbes that are more consistent across replicate individuals. Our data support this hypothesis, with the composition of bacterial (16S rRNA gene) and microeukaryote (18S rRNA gene) communities showing significant differences across the four anatomical regions, with the surfaces of older tissues (holdfast and blade tips) harboring significantly greater microbial richness compared to the younger tissues of the meristematic region. Additional samples collected from the surfaces of new L. setchellii recruits (<1y old) also showed differences in microbial community structure across anatomical regions, which demonstrates that these microbial differences are established early. We also observed this pattern in two additional algal species, suggesting that microbial community structure across host anatomy may be a common feature of the seaweed microbiome.
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Kelp , Laminaria , Microbiota , Bacterias/genética , ARN Ribosómico 16S/genéticaRESUMEN
Ethylene-forming enzyme (EFE) is an ambifunctional iron(II)- and 2-oxoglutarate-dependent (Fe/2OG) oxygenase. In its major (EF) reaction, it converts carbons 1, 2, and 5 of 2OG to CO2 and carbons 3 and 4 to ethylene, a four-electron oxidation drastically different from the simpler decarboxylation of 2OG to succinate mediated by all other Fe/2OG enzymes. EFE also catalyzes a minor reaction, in which the normal decarboxylation is coupled to oxidation of l-arginine (a required activator for the EF pathway), resulting in its conversion to l-glutamate semialdehyde and guanidine. Here we show that, consistent with precedent, the l-Arg-oxidation (RO) pathway proceeds via an iron(IV)-oxo (ferryl) intermediate. Use of 5,5-[2H2]-l-Arg slows decay of the ferryl complex by >16-fold, implying that RO is initiated by hydrogen-atom transfer (HAT) from C5. That this large substrate deuterium kinetic isotope effect has no impact on the EF:RO partition ratio implies that the same ferryl intermediate cannot be on the EF pathway; the pathways must diverge earlier. Consistent with this conclusion, the variant enzyme bearing the Asp191Glu ligand substitution accumulates â¼4 times as much of the ferryl complex as the wild-type enzyme and exhibits a â¼40-fold diminished EF:RO partition ratio. The selective detriment of this nearly conservative substitution to the EF pathway implies that it has unusually stringent stereoelectronic requirements. An active-site, like-charge guanidinium pair, which involves the l-Arg substrate/activator and is unique to EFE among four crystallographically characterized l-Arg-modifying Fe/2OG oxygenases, may serve to selectively stabilize the transition state leading to the unique EF branch.
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Arginina/química , Hierro/química , Ácidos Cetoglutáricos/metabolismo , Oxigenasas/metabolismo , Modelos Moleculares , Oxidación-Reducción , Oxigenasas/química , Conformación ProteicaRESUMEN
(S)-2-Hydroxypropylphosphonate [(S)-2-HPP, 1] epoxidase (HppE) reduces H2O2 at its nonheme-iron cofactor to install the oxirane "warhead" of the antibiotic fosfomycin. The net replacement of the C1 pro-R hydrogen of 1 by its C2 oxygen, with inversion of configuration at C1, yields the cis-epoxide of the drug [(1R,2S)-epoxypropylphosphonic acid (cis-Fos, 2)]. Here we show that HppE achieves â¼95% selectivity for C1 inversion and cis-epoxide formation via steric guidance of a radical-coupling mechanism. Published structures of the HppE·FeII·1 and HppE·ZnII·2 complexes reveal distinct pockets for C3 of the substrate and product and identify four hydrophobic residues-Leu120, Leu144, Phe182, and Leu193-close to C3 in one of the complexes. Replacement of Leu193 in the substrate C3 pocket with the bulkier Phe enhances stereoselectivity (cis:trans â¼99:1), whereas the Leu120Phe substitution in the product C3 pocket diminishes it (â¼82:18). Retention of C1 configuration and trans-epoxide formation become predominant with the bulk-reducing Phe182Ala substitution in the substrate C3 pocket (â¼13:87), trifluorination of C3 (â¼23:77), or both (â¼1:99). The effect of C3 trifluorination is counteracted by the more constrained substrate C3 pockets in the Leu193Phe (â¼56:44) and Leu144Phe/Leu193Phe (â¼90:10) variants. The ability of HppE to epoxidize substrate analogues bearing halogens at C3, C1, or both is inconsistent with a published hypothesis of polar cyclization via a C1 carbocation. Rather, specific enzyme-substrate contacts drive inversion of the C1 radical-as proposed in a recent computational study-to direct formation of the more potently antibacterial cis-epoxide by radicaloid C-O coupling.
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Compuestos Epoxi/metabolismo , Fosfomicina/biosíntesis , Oxidorreductasas/metabolismo , Compuestos Epoxi/química , Fosfomicina/química , Radicales Libres/química , Radicales Libres/metabolismo , Conformación Molecular , Oxidorreductasas/química , Oxidorreductasas/genética , EstereoisomerismoRESUMEN
Iron(II)- and 2-(oxo)-glutarate-dependent (Fe/2OG) oxygenases catalyze a diverse array of oxidation reactions via a common iron(IV)-oxo (ferryl) intermediate. Although the intermediate has been characterized spectroscopically, its short lifetime has precluded crystallograhic characterization. In solution, the ferryl was first observed directly in the archetypal Fe/2OG hydroxylase, taurine:2OG dioxygenase (TauD). Here, we substitute the iron cofactor of TauD with the stable vanadium(IV)-oxo (vanadyl) ion to obtain crystal structures mimicking the key ferryl complex. Intriguingly, whereas the structure of the TauD·(VIV-oxo)·succinate·taurine complex exhibits the expected orientation of the V≡O bond-trans to the His255 ligand and toward the C-H bond to be cleaved, in what has been termed the in-line configuration-the TauD·(VIV-oxo) binary complex is best modeled with its oxo ligand trans to Asp101. This off-line-like configuration is similar to one recently posited as a means to avoid hydroxylation in Fe/2OG enzymes that direct other outcomes, though neither has been visualized in an Fe/2OG structure to date. Whereas an off-line (trans to the proximal His) or off-line-like (trans to the carboxylate ligand) ferryl is unlikely to be important in the hydroxylation reaction of TauD, the observation that the ferryl may deviate from an in-line orientation in the absence of the primary substrate may explain the enzyme's mysterious self-hydroxylation behavior, should the oxo ligand lie trans to His99. This finding reinforces the potential for analogous functional off-line oxo configurations in halogenases, desaturases, and/or cyclases.
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Hierro/química , Oxigenasas de Función Mixta/química , Imitación Molecular , Vanadatos/química , Dominio Catalítico , Cristalización , Cristalografía por Rayos X , Escherichia coli/química , Enlace de Hidrógeno , Hidroxilación , Oxigenasas de Función Mixta/aislamiento & purificación , Estructura Secundaria de Proteína , Ácido Succínico/química , Taurina/química , Espectroscopía de Absorción de Rayos XRESUMEN
Indoleamine 2,3-dioxygenase (IDO1) is a heme enzyme that catalyzes the oxygenation of the indole ring of tryptophan to afford N-formylkynurenine. This activity significantly suppresses the immune response, mediating inflammation and autoimmune reactions. These consequential effects are regulated through redox changes in the heme cofactor of IDO1, which autoxidizes to the inactive ferric state during turnover. This change in redox status increases the lability of the heme cofactor leading to further suppression of activity. The cell can thus regulate IDO1 activity through the supply of heme and reducing agents. We show here that polysulfides bind to inactive ferric IDO1 and reduce it to the oxygen-binding ferrous state, thus activating IDO1 to maximal turnover even at low, physiologically significant concentrations. The on-rate for hydrogen disulfide binding to ferric IDO1 was found to be >106 M-1 s-1 at pH 7 using stopped-flow spectrometry. Fe K-edge XANES and EPR spectroscopy indicated initial formation of a low-spin ferric sulfur-bound species followed by reduction to the ferrous state. The µM affinity of polysulfides for IDO1 implicates these polysulfides as important signaling factors in immune regulation through the kynurenine pathway. Tryptophan significantly enhanced the relatively lower-affinity binding of hydrogen sulfide to IDO1, inspiring the use of the small molecule 3-mercaptoindole (3MI), which selectively binds to and activates ferric IDO1. 3MI sustains turnover by catalytically transferring reducing equivalents from glutathione to IDO1, representing a novel strategy of upregulating innate immunosuppression for treatment of autoimmune disorders. Reactive sulfur species are thus likely unrecognized immune-mediators with potential as therapeutic agents through these interactions with IDO1.