The interplay between metal ion binding and the activity of thiol proteins, particularly within the protein disulfide isomerase family, remains an area of active investigation due to the critical role that these proteins play in many vital processes. This research investigates the interaction between recombinant human PDIA1 and zinc ions, focusing on the subsequent implications for PDIA1\'s conformational stability and enzymatic activity. Employing isothermal titration calorimetry and differential scanning calorimetry, we systematically compared the zinc binding capabilities of both oxidized and reduced forms of PDIA1 and assessed the structural consequences of this interaction. Our results demonstrate that PDIA1 can bind zinc both in reduced and oxidized states, but with significantly different stoichiometry and more pronounced conformational effects in the reduced form of PDIA1. Furthermore, zinc binding was observed to inhibit the catalytic activity of reduced-PDIA1, likely due to induced alterations in its conformation. These findings unveil a potential regulatory mechanism in PDIA1, wherein metal ion binding under reductive conditions modulates its activity. Our study highlights the potential role of zinc in regulating the catalytic function of PDIA1 through conformational modulation, suggesting a nuanced interplay between metal binding and protein stability in the broader context of cellular redox regulation.

Hydropersulfides (RSSH) are oxidized thiol (RSH) derivatives that have been shown to be biologically prevalent with likely important functions (along with other polysulfur compounds). The functional utility of RSSH can be gleaned from their unique chemical properties. That is, RSSH possess chemical reactivity not present in other biologically relevant sulfur species that should allow them to be used in specific ways in biology as effector/signaling molecules. For example, compared to RSH, RSSH are considered to be superior nucleophiles, reductants and metal ligands. Moreover, unlike RSH, RSSH can be either reductants/nucleophiles or oxidants/electrophiles depending on the protonated state. It has also become clear that studies related to the chemical biology and physiology of hydrogen suflide (H2S) must also consider the effects of RSSH (and related polysulfur species) as they are biochemically linked. Herein is a discussion of the relevant chemistry of RSSH that can serve as a basis for understanding how RSSH can be used by cells to, for example, combat stresses and used in signaling. Also, discussed are some current experimental studies regarding the biological activity of RSSH that can be explained by their chemical properties.

Allelochemicals are considered an environment-friendly and promising alternative for weed management, although much effort is still needed for understanding their mode of action and then promoting their use in plant allelopathy management practices. Here, we report that Inuloxin A (InA), an allelochemical isolated from Dittrichia viscosa, inhibited root elongation and growth of seedlings of Lycopersicon esculentum and Lepidium sativum at the highest concentrations tested. InA-induced antioxidant responses in the seedlings were investigated by analysing the contents of glutathione (GSH) and ascorbate (ASC), and their oxidized forms, dehydroascorbate (DHA), and glutathione disulphide (GSSG), as well as the redox state of thiol-containing proteins. An increase in ASC, DHA, and GSH levels at high concentrations of InA, after 3 and 6 days, were observed. Moreover, the ASC/DHA + ASC and GSH/GSSG + GSH ratios showed a shift towards the oxidized form. Our study provides the first insight into how the cell redox system responds and adapts to InA phytotoxicity, providing a framework for further molecular studies.

Significance: Hydrogen sulfide (H2S) is reported to be an important mediator involved in numerous physiological processes. H2S and hydropersulfides (RSSH) species are intimately linked biochemically. Therefore, interest in the mechanisms of the biological activity of H2S has led to investigations of the chemical biology of RSSH since they are likely to coexist in a biological system. Currently it is hypothesized that RSSH may be responsible for a least part of the observed H2S-mediated biology/physiology. Recent Advances: It has been recently touted that thiols (RSH) and RSSH have some important differences in terms of their chemical biology and that the generation of RSSH from RSH is purposeful to exploit these chemical differences as a response to a physiological or biological stress. This transformation may represent an unappreciated/unrecognized biological mechanism for dealing with cellular stresses. Critical Issues: Although recent studies indicate a diverse and potentially important chemical biology associated with RSSH species, these ideas have their foundations in early studies (some over 60 years old). It is vital to recognize the nature of this early work to fully appreciate the current ideas regarding RSSH biology. Importantly, these early studies were performed before the realization of purposeful H2S biosynthesis (before 1996). Future Directions: Taking clues from the past studies of RSSH chemistry and biology, progress in delineating the chemical biology of RSSH will continue. Determination of the possible relevance of RSSH chemical biology to signaling and cellular physiology will be a primary focus of many future studies.

Redox modification, a post-translational modification, has been demonstrated to be significant for many physiological pathways and biological processes in both eukaryotes and prokaryotes. However, little is known about the global profile of protein redox modification in fungi. To explore the roles of redox modification in the plant pathogenic fungi, a global thiol proteome survey was performed in the model fungal pathogen Magnaporthe oryzae. A total of 3713 redox modification sites from 1899 proteins were identified through a mix sample containing mycelia with or without oxidative stress, conidia, appressoria, and invasive hyphae of M. oryzae. The identified thiol-modified proteins were performed with protein domain, subcellular localization, functional classification, metabolic pathways, and protein-protein interaction network analyses, indicating that redox modification is associated with a wide range of biological and cellular functions. These results suggested that redox modification plays important roles in fungal growth, conidium formation, appressorium formation, as well as invasive growth. Interestingly, a large number of pathogenesis-related proteins were redox modification targets, suggesting the significant roles of redox modification in pathogenicity of M. oryzae. This work provides a global insight into the redox proteome of the pathogenic fungi, which built a groundwork and valuable resource for future studies of redox modification in fungi.

Significance: Supracellular redox networks regulating cell-extracellular matrix (ECM) and organ system architecture merge with structural and functional (catalytic or allosteric) properties of disulfide bonds. This review addresses emerging evidence that exported thiol oxidoreductases (TORs), such as thioredoxin, protein disulfide isomerases (PDIs), quiescin sulfhydryl oxidases (QSOX)1, and peroxiredoxins, composing a peri/epicellular (pec)TOR pool, mediate relevant signaling. pecTOR functions depend mainly on kinetic and spatial regulation of thiol-disulfide exchange reactions governed by redox potentials, which are modulated by exported intracellular low-molecular-weight thiols, together conferring signal specificity. Recent Advances: pecTOR redox-modulates several targets including integrins, ECM proteins, surface molecules, and plasma components, although clear-cut documentation of direct effects is lacking in many cases. TOR catalytic pathways, displaying common patterns, culminate in substrate thiol reduction, oxidation, or isomerization. Peroxiredoxins act as redox/peroxide sensors, contrary to PDIs, which are likely substrate-targeted redox modulators. Emerging evidence suggests important pecTOR roles in patho(physio)logical processes, including blood coagulation, vascular remodeling, mechanosensing, endothelial function, immune responses, and inflammation. Critical Issues: Effects of pecPDIs supporting thrombosis/platelet activation have been well documented and reached the clinical arena. Roles of pecPDIA1 in vascular remodeling/mechanosensing are also emerging. Extracellular thioredoxin and pecPDIs redox-regulate immunoinflammation. Routes of TOR externalization remain elusive and appear to involve Golgi-independent routes. pecTORs are particularly accessible drug targets. Future Directions: Further understanding mechanisms of thiol redox reactions and developing assays for assessing pecTOR redox activities remain important research avenues. Also, addressing pecTORs as disease markers and achieving more efficient/specific drugs for pecTOR modulation are major perspectives for diagnostic/therapeutic improvements.

Redox-related plasma proteins are candidate reporters of protein signatures associated with endothelial structure/function. Thiol-proteins from protein disulfide isomerase (PDI) family are unexplored in this context. Here, we investigate the occurrence and physiological significance of a circulating pool of PDI in healthy humans. We validated an assay for detecting PDI in plasma of healthy individuals. Our results indicate high inter-individual (median = 330 pg/mL) but low intra-individual variability over time and repeated measurements. Remarkably, plasma PDI levels could discriminate between distinct plasma proteome signatures, with PDI-rich (>median) plasma differentially expressing proteins related to cell differentiation, protein processing, housekeeping functions and others, while PDI-poor plasma differentially displayed proteins associated with coagulation, inflammatory responses and immunoactivation. Platelet function was similar among individuals with PDI-rich vs. PDI-poor plasma. Remarkably, such protein signatures closely correlated with endothelial function and phenotype, since cultured endothelial cells incubated with PDI-poor or PDI-rich plasma recapitulated gene expression and secretome patterns in line with their corresponding plasma signatures. Furthermore, such signatures translated into functional responses, with PDI-poor plasma promoting impairment of endothelial adhesion to fibronectin and a disturbed pattern of wound-associated migration and recovery area. Patients with cardiovascular events had lower PDI levels vs. healthy individuals. This is the first study describing PDI levels as reporters of specific plasma proteome signatures directly promoting contrasting endothelial phenotypes and functional responses.

Thiol groups are crucially involved in signaling/homeostasis through oxidation, reduction, and disulphide exchange. The overall thiol pool is the resultant of several individual pools of small compounds (e.g. cysteine), peptides (e.g. glutathione), and thiol proteins (e.g. thioredoxin (Trx)), which are not in equilibrium and present specific oxidized/reduced ratios. This review addresses mechanisms and implications of circulating plasma thiol/disulphide redox pools, which are involved in several physiologic processes and explored as disease biomarkers. Thiol pools are regulated by mechanisms linked to their intrinsic reactivity against oxidants, concentration of antioxidants, thiol-disulphide exchange rates, and their dynamic release/removal from plasma. Major thiol couples determining plasma redox potential (Eh) are reduced cysteine (CyS)/cystine (the disulphide form of cysteine) (CySS), followed by GSH/disulphide-oxidized glutathione (GSSG). Hydrogen peroxide and hypohalous acids are the main plasma oxidants, while water-soluble and lipid-soluble small molecules are the main antioxidants. The thiol proteome and thiol-oxidoreductases are emerging investigative areas given their specific disease-related responses (e.g. protein disulphide isomerases (PDIs) in thrombosis). Plasma cysteine and glutathione redox couples exhibit pro-oxidant changes directly correlated with ageing/age-related diseases. We further discuss changes in thiol-disulphide redox state in specific groups of diseases: cardiovascular, cancer, and neurodegenerative. These results indicate association with the disease states, although not yet clear-cut to yield specific biomarkers. We also highlight mechanisms whereby thiol pools affect atherosclerosis pathophysiology. Overall, it is unlikely that a single measurement provides global assessment of plasma oxidative stress. Rather, assessment of individual thiol pools and thiol-proteins specific to any given condition has more solid and logical perspective to yield novel relevant information on disease risk and prognosis.

Protein disulfide isomerases are thiol oxidoreductase chaperones from thioredoxin superfamily. As redox folding catalysts from the endoplasmic reticulum (ER), their roles in ER-related redox homeostasis and signaling are well-studied. PDIA1 exerts thiol oxidation/reduction and isomerization, plus chaperone effects. Also, substantial evidence indicates that PDIs regulate thiol-disulfide switches in other cell locations such as cell surface and possibly cytosol. Subcellular PDI translocation routes remain unclear and seem Golgi-independent. The list of signaling and structural proteins reportedly regulated by PDIs keeps growing, via thiol switches involving oxidation, reduction and isomerization, S-(de)nytrosylation, (de)glutathyonylation and protein oligomerization. PDIA1 is required for agonist-triggered Nox NADPH oxidase activation and cell migration in vascular cells and macrophages, while PDIA1-dependent cytoskeletal regulation appears a converging pathway. Extracellularly, PDIs crucially regulate thiol redox signaling of thrombosis/platelet activation, e.g., integrins, and PDIA1 supports expansive caliber remodeling during injury repair via matrix/cytoskeletal organization. Some proteins display regulatory PDI-like motifs. PDI effects are orchestrated by expression levels or post-translational modifications. PDI is redox-sensitive, although probably not a mass-effect redox sensor due to kinetic constraints. Rather, the \"all-in-one\" organization of its peculiar redox/chaperone properties likely provide PDIs with precision and versatility in redox signaling, making them promising therapeutic targets.

Nitroxyl (HNO) possesses unique and potentially important biological/physiological activity that is currently mechanistically ill-defined. Previous work has shown that the likely biological targets for HNO are thiol proteins, oxidized metalloproteins (i.e. ferric heme proteins) and, most likely, selenoproteins. Interestingly, these are the same classes of proteins that interact with H2O2. In fact, these classes of proteins not only react with H2O2, and thus potentially responsible for the signaling actions of H2O2, but are also responsible for the degradation of H2O2. Therefore, it is not unreasonable to speculate that HNO can affect H2O2 degradation by interacting with H2O2-degrading proteins possibly leading to an increase in H2O2-mediated signaling. Moreover, considering the commonality between HNO and H2O2 biological targets, it also seems likely that HNO-mediated signaling can also be due to reactivity at otherwise H2O2-reactive sites. Herein, it is found that HNO does indeed inhibit H2O2 degradation via inhibition of H2O2-metaboilizing proteins. Also, it is found that in a system known to be regulated by H2O2 (T cell activation), HNO behaves similarly to H2O2, indicating that HNO- and H2O2-signaling may be similar and/or intimately related.