Reassessing enzyme kinetics Ferrall-Fairbanks, Meghan C.; Kieslich, Chris A.; Platt, Manu O.
Proceedings of the National Academy of Sciences - PNAS,
02/2020, Letnik:
117, Številka:
6
Journal Article
Recenzirano
Enzymes are catalysts in biochemical reactions that, by definition, increase rates of reactions without being altered or destroyed. However, when that enzyme is a protease, a subclass of enzymes that ...hydrolyze other proteins, and that protease is in a multiprotease system, protease-as-substrate dynamics must be included, challenging assumptions of enzyme inertness, shifting kinetic predictions of that system. Protease-on-protease inactivating hydrolysis can alter predicted protease concentrations used to determine pharmaceutical dosing strategies. Cysteine cathepsins are proteases capable of cathepsin cannibalism, where one cathepsin hydrolyzes another with substrate present, and misunderstanding of these dynamics may causemiscalculations of multiple proteases working in one proteolytic network of interactions occurring in a defined compartment. Once rates for individual protease-on-protease binding and catalysis are determined, proteolytic network dynamics can be explored using computational models of cooperative/competitive degradation by multiple proteases in one system, while simultaneously incorporating substrate cleavage. During parameter optimization, it was revealed that additional distraction reactions, where inactivated proteases become competitive inhibitors to remaining, active proteases, occurred, introducing another network reaction node. Taken together, improved predictions of substrate degradation in a multiple protease network were achieved after including reaction terms of autodigestion, inactivation, cannibalism, and distraction, altering kinetic considerations from other enzymatic systems, since enzyme can be lost to proteolytic degradation. We compiled and encoded these dynamics into an online platform (https://plattlab.shinyapps.io/catKLS/) for individual users to test hypotheses of specific perturbations to multiple cathepsins, substrates, and inhibitors, and predict shifts in proteolytic network reactions and system dynamics.
Cysteine cathepsins are lysosomal peptidases involved on one hand in general intracellular protein degradation and, on the other, in the regulation of a number of specific physiological processes. ...Their integral role in extracellular matrix degradation and in processing growth factors, hormones and adhesion proteins is a driving force in cancer progression, triggering tumor proliferation, invasion, angiogenesis and metastasis. Cancer stem cells are proposed to be a main factor of tumor initialization, heterogeneity and resistance to therapy. Recent studies have uncovered increased expression, aberrant localization and disturbed functions of certain cysteine cathepsins in these cells, in particular in glioblastoma stem cells. Here, we review the research that have underlined the expression patterns and roles of cysteine cathepsins in cancer stem cells, and emphasize the involvement of cysteine cathepsins in caspase-independent cell death and in regulating interaction between cancer stem cells and immune cells such as natural killer cells.
Unlike other fluoride-based caries preventive agents, silver diamine fluoride (SDF) can simultaneously prevent and arrest coronal and root dentine caries. The profound clinical success of SDF has ...drawn many clinicians and researchers to study the mechanism of SDF in arresting dentine caries. This critical review discusses how silver and fluoride contribute to caries arrest, in terms of their effects on bacteria as well as on the mineral and organic content of dentine. Silver interacts with bacterial cell membrane and bacterial enzymes, which can inhibit bacterial growth. Silver can also dope into hydroxyapatite and have an antibacterial effect on silver-doped hydroxyapatite. Furthermore, silver is also a strong inhibitor of cathepsins and inhibits dentine collagen degradation. Early studies proposed that silver hardened caries lesions by forming silver phosphate. However, recent studies found that little silver phosphate remained on the arrested dentine lesion. The principal silver precipitate was silver chloride, which could not contribute to the significant hardening of the arrested lesions. On the other hand, fluoride enhances mineral formation by forming fluorohydroxyapatite with reduced solubility. A significant increase in microhardness occurs with an elevated level of calcium and phosphorus but not silver on the surface layer of the arrested dentine caries lesion following SDF treatment. Fluoride also inhibits matrix metalloproteinases activities and therefore inhibits dentine collagen degradation. The combination of silver and fluoride in an alkaline solution has a synergistic effect in arresting dentine caries. The alkaline property of SDF provides an unfavorable environment for collagen enzyme activation. Understanding the mechanisms of SDF in arresting dentine caries helps clinicians to develop appropriate protocols for the use of SDF in clinical care.
Cysteine cathepsins are key regulators of the innate and adaptive arms of the immune system. Their expression, activity, and subcellular localization are associated with the distinct development and ...differentiation stages of immune cells. They promote the activation of innate myeloid immune cells since they contribute to toll-like receptor signaling and to cytokine secretion. Furthermore, they control lysosomal biogenesis and autophagic flux, thus affecting innate immune cell survival and polarization. They also regulate bidirectional communication between the cell exterior and the cytoskeleton, thus influencing cell interactions, morphology, and motility. Importantly, cysteine cathepsins contribute to the priming of adaptive immune cells by controlling antigen presentation and are involved in cytotoxic granule mediated killing in cytotoxic T lymphocytes and natural killer cells. Cathepins'aberrant activity can be prevented by their endogenous inhibitors, cystatins. However, dysregulated proteolysis contributes significantly to tumor progression also by modulation of the antitumor immune response. Especially tumor-associated myeloid cells, such as tumor-associated macrophages and myeloid-derived suppressor cells, which are known for their tumor promoting and immunosuppressive functions, constitute the major source of excessive cysteine cathepsin activity in cancer. Since they are enriched in the tumor microenvironment, cysteine cathepsins represent exciting targets for development of new diagnostic and therapeutic moieties.
Lysosomal cysteine cathepsins are a family of proteases that are involved in a myriad of cellular processes from proteolytic degradation in the lysosome to bone resorption. These proteins mature ...following the cleavage of a pro-domain in the lysosome to become either exo- or endo-peptidases. The cathepsins B, C, L, S and Z have been implicated in NLRP3 inflammasome activation following their activation with ATP, monosodium urate, silica crystals, or bacterial components, among others. These five cathepsins have both compensatory and independent functions in NLRP3 inflammasome activation. There is much evidence in the literature to support the release of cathepsin B following lysosomal membrane degradation which leads to NLRP3 inflammasome activation. This is likely due to a hitherto unidentified role of this protein in the cytoplasm, although other interactions with autophagy proteins and within lysosomes have been proposed. Cathepsin C is involved in the processing of neutrophil IL-1β through processing of upstream proteases. Cathepsin Z is non-redundantly required for NLRP3 inflammasome activation following nigericin, ATP and monosodium urate activation. Lysosomal cysteine cathepsins are members of a diverse and complementary family, and likely share both overlapping and independent functions in NLRP3 inflammasome activation.
•Multiple cathepsins are involved in NLRP3 inflammasome activation; the context of activation may determine which cathepsins—or whether cathepsins—are involved.•Cathepsins B, C, L, S, Z may share overlapping or compensatory functions in NLRP3 inflammasome activation.•Cathepsin B has been implicated in inflammasome activation using compounds that induce lysosomal membrane permeabilization and may involve negative regulation of autophagy.•Cathepsin C has been reported to activate neutrophil serine proteases which cleave IL-1β independently of caspase-1.•Cathepsin Z is involved in NLRP3 inflammasome activation independently of lysosomal membrane permeabilization.
Cysteine cathepsins have been for a long time considered to execute mainly nonspecific bulk proteolysis in the endolysosomal system. However, this view has been changing profoundly over the last ...decade as cathepsins were found in the cytoplasm, nucleus and in the extracellular milieu. Cathepsins are currently gaining increased attention largely because of their extracellular roles associated with disease development and progression. While kept under tight control under physiological conditions, their dysregulated and elevated activity in the extracellular milieu are distinctive hallmarks of numerous diseases such as various cancers, inflammatory disorders, rheumatoid arthritis, bone disorders and heart diseases. In this review, we discuss cysteine cathepsins with a major focus on their extracellular roles and extracellular proteolytic targets beyond degradation of the extracellular matrix. We further highlight the perspectives of cathepsin research and novel avenues in cathepsin-based diagnostic and therapeutic applications.
•Cysteine cathepsins are often highly upregulated in inflammation-associated diseases linked with ECM remodelling.•Proteomic studies led to identification of several new cathepsin substrates revealing novel roles of cathepsins in the ECM.•Cathepsin K inhibitor Odanacatib failed in advanced clinical trials, but alternatives seem to be on a way.•Diagnostic imaging of cysteine cathepsins in oncology has entered clinical trials•Cathepsins are largely used as activators of prodrugs and antibody-drug conjugates in oncology
Cysteine cathepsins are primarily involved in the degradation and recycling of proteins in endo-lysosomal compartments but are also gaining recognition as pivotal proteolytic contributors to various ...immune functions. Through their extracellular proteolytic activities within the hematopoietic stem cell niche, they are involved in progenitor cell mobilization and differentiation. Cysteine cathepsins, such as cathepsins L and S contribute to antigen-induced adaptive immunity through major histocompatibility complex class II antigen presentation whereas cathepsin X regulates T-cell migration. By regulating toll-like receptor signaling and cytokine secretion cysteine cathepsins activate innate immune cells and affect their functional differentiation. Cathepsins C and H are expressed in cytotoxic T lymphocytes and natural killer cells and are involved in processing of pro-granzymes into proteolytically active forms. Cytoplasmic activities of cathepsins B and L contribute to the maintenance of homeostasis of the adaptive immune response by regulating cell death of T and B lymphocytes. The expression pattern, localization, and activity of cysteine cathepsins is tightly connected to their function in immune cells. Furthermore, cysteine cathepsins together with their endogenous inhibitors, serve as mediators in the interplay between cancer and immune cells that results in immune cell anergy. The aim of the present article is to review the mechanisms of dysregulation of cysteine cathepsins and their inhibitors in relation to immune dysfunction to address new possibilities for regulation of their function.
Biomedical research often focuses on properties that differentiate between diseased and healthy tissue; one of the current focuses is elevated expression and altered localisation of proteases. Among ...these proteases, dysregulation of cysteine cathepsins can frequently be observed in inflammation-associated diseases, which tips the functional balance from normal physiological to pathological manifestations. Their overexpression and secretion regularly exhibit a strong correlation with the development and progression of such diseases, making them attractive pharmacological targets. But beyond their mostly detrimental role in inflammation-associated diseases, cysteine cathepsins are physiologically highly important enzymes involved in various biological processes crucial for maintaining homeostasis and responding to different stimuli. Consequently, several challenges have emerged during the efforts made to translate basic research data into clinical applications. In this review, we present both physiological and pathological roles of cysteine cathepsins and discuss the clinical potential of cysteine cathepsin-targeting strategies for disease management and diagnosis.
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Proanthocyanidin-rich plant-derived agents have been shown to enhance dentin biomechanical properties and resistance to collagenase degradation. This study systematically investigated ...the interaction of chemically well-defined monomeric catechins with dentin extracellular matrix components by evaluating dentin mechanical properties as well as activities of matrix metalloproteinases (MMPs) and cysteine-cathepsins (CTs). Demineralized dentin beams (n=15) were incubated for 1h with 0.65% (+)-catechin (C), (−)-catechin gallate (CG), (−)-gallocatechin gallate (GCG), (−)-epicatechin (EC), (−)-epicatechin gallate (ECG), (−)-epigallocatechin (EGC) and (−)-epigallocatechin-3-gallate (EGCG). The modulus of elasticity (E) and the fold increase in E were determined by comparing specimens at baseline and after treatment. Biodegradation rates were assessed by differences in percentage of dry mass before and after incubation with bacterial collagenase. The inhibition of MMP-9 and CT-B by 0.65, 0.065 and 0.0065% of each catechin was determined using fluorimetric proteolytic assay kits. All monomeric catechins led to a significant increase in E. EGCG showed the highest fold increase in E, followed by ECG, CG and GCG. EGCG, ECG, GCG and CG significantly lowered biodegradation rates and inhibited both MMP-9 and CT-B at a concentration of 0.65%. Overall, the 3-O-galloylated monomeric catechins are clearly more potent than their non-galloylated analogues in improving dentin mechanical properties, stabilizing collagen against proteolytic degradation, and inhibiting the activity of MMPs and CTs. The results indicate that galloylation is a key pharmacophore in the monomeric and likely also in the oligomeric proanthocyanidins that exhibit high cross-linking potential for dentin extracellular matrix.
Papain-like cysteine peptidases are a diverse family of peptidases found in most known organisms. In eukaryotes, they are divided into multiple evolutionary groups, which can be clearly distinguished ...on the basis of the structural characteristics of the proenzymes. Most of them are endopeptidases; some, however, evolved into exopeptidases by obtaining additional structural elements that restrict the binding of substrate into the active site. In humans, papain-like peptidases, also called cysteine cathepsins, act both as non-specific hydrolases and as specific processing enzymes. They are involved in numerous physiological processes, such as antigen presentation, extracellular matrix remodeling, and hormone processing. Their activity is tightly regulated and dysregulation of one or more cysteine cathepsins can result in severe pathological conditions, such as cardiovascular diseases and cancer. Other organisms can utilize papain-like peptidases for different purposes and they are often part of host-pathogen interactions. Numerous parasites, such as
and flukes, utilize papain-like peptidases for host invasion, whereas plants, in contrast, use these enzymes for host defense. This review presents a state-of-the-art description of the structure and phylogeny of papain-like peptidases as well as an overview of their physiological and pathological functions in humans and in other organisms.
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