Profiling of coronaviral M and enteroviral 3C specificity provides a framework for the development of broad-spectrum antiviral compounds.

The main protease from coronaviruses and the 3C protease from enteroviruses play a crucial role in processing viral polyproteins, making them attractive targets for the development of antiviral agents. In this study, we employed a combinatorial chemistry approach-HyCoSuL-to compare the substrate specificity profiles of the main and 3C proteases from alphacoronaviruses, betacoronaviruses, and enteroviruses. The obtained data demonstrate that coronavirus Ms exhibit overlapping substrate specificity in all binding pockets, whereas the 3C from enterovirus displays slightly different preferences toward natural and unnatural amino acids at the P4-P2 positions.

Substrate specificity profiling of heat-sensitive serine protease from the fungus Onygena corvina.

Proteases catalyze hydrolysis of amide bonds within peptides and proteins, therefore they play crucial functions for organism functioning, but also in industry to facilitate numerous processes. Feather-degrading fungus Onygena corvina (O. corvina) is loaded with numerous proteases that can be utilized for variety of applications.

Targeting Microglial Immunoproteasome: A Novel Approach in Neuroinflammatory-Related Disorders.

It is widely acknowledged that the aging process is linked to the accumulation of damaged and misfolded proteins. This phenomenon is accompanied by a decrease in proteasome (c20S) activity, concomitant with an increase in immunoproteasome (i20S) activity. These changes can be attributed, in part, to the chronic neuroinflammation that occurs in brain tissues.

Chemical tools to define and manipulate interferon-inducible Ubl protease USP18.

Ubiquitin-specific protease 18 (USP18) is a multifunctional cysteine protease primarily responsible for deconjugating interferon-inducible ubiquitin-like (Ubl) modifier ISG15 from protein substrates. Here, we report the design and synthesis of activity-based probes (ABPs) capable of selectively detecting USP18 activity over other ISG15 cross-reactive deubiquitinases (DUBs) by incorporating unnatural amino acids into the C-terminal tail of ISG15. Combining with a ubiquitin-based DUB ABP, the selective USP18 ABP is employed in a chemoproteomic screening platform to identify and assess inhibitors of DUBs including USP18.

Mapping the substrate-binding subsite specificity of a Porphyromonas gingivalis Tpr peptidase.

Calcium-dependent peptidases of the calpain family are widespread in eukaryotes but uncommon in prokaryotes. A few bacterial calpain homologs have been discovered but none of them have been characterized in detail. Here we present an in-depth substrate specificity analysis of the bacterial calpain-like peptidase Tpr from Porphyromonas gingivalis.

Ebselen derivatives inhibit SARS-CoV-2 replication by inhibition of its essential proteins: PL and M proteases, and nsp14 guanine N7-methyltransferase.

Proteases encoded by SARS-CoV-2 constitute a promising target for new therapies against COVID-19. SARS-CoV-2 main protease (M, 3CL) and papain-like protease (PL) are responsible for viral polyprotein cleavage-a process crucial for viral survival and replication. Recently it was shown that 2-phenylbenzisoselenazol-3(2H)-one (ebselen), an organoselenium anti-inflammatory small-molecule drug, is a potent, covalent inhibitor of both the proteases and its potency was evaluated in enzymatic and antiviral assays.

SARS-CoV-2 M Protease Variants of Concern Display Altered Viral Substrate and Cell Host Target Galectin-8 Processing but Retain Sensitivity toward Antivirals.

The main protease of SARS-CoV-2 (M) is the most promising drug target against coronaviruses due to its essential role in virus replication. With newly emerging variants there is a concern that mutations in M may alter the structural and functional properties of protease and subsequently the potency of existing and potential antivirals. We explored the effect of 31 mutations belonging to 5 variants of concern (VOCs) on catalytic parameters and substrate specificity, which revealed changes in substrate binding and the rate of cleavage of a viral peptide.

Imaging of proteases using activity-based probes.

Proteases (proteolytic enzymes) are proteins that catalyze one of the most important biochemical reactions, namely the hydrolysis of the peptide bond in peptide and protein substrates. Therefore these molecular biocatalysts participate in virtually all living processes. The proper balance between intact and processed protease substrates enables to maintenance of homeostasis from a single-cell level to the whole living system.

Fluorescent Activity-Based Probe To Image and Inhibit Factor XIa Activity in Human Plasma.

Anticoagulation therapy is a mainstay of the treatment of thrombotic disorders; however, conventional anticoagulants trade antithrombotic benefits for bleeding risk. Factor (f) XI deficiency, known as hemophilia C, rarely causes spontaneous bleeding, suggesting that fXI plays a limited role in hemostasis. In contrast, individuals with congenital fXI deficiency display a reduced incidence of ischemic stroke and venous thromboembolism, indicating that fXI plays a role in thrombosis.

Investigation of osteoclast cathepsin K activity in osteoclastogenesis and bone loss using a set of chemical reagents.

Cathepsin K (CatK) is a lysosomal cysteine protease whose highest expression is found in osteoclasts, which are the cells responsible for bone resorption. Investigations of the functions and physiological relevance of CatK have often relied on antibody-related techniques, which makes studying its activity patterns a challenging task. Hence, we developed a set of chemical tools for the investigation of CatK activity.

Amino acid variants of SARS-CoV-2 papain-like protease have impact on drug binding.

The novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused both a health and economic crisis around the world. Its papain-like protease (PLpro) is one of the protein targets utilized in designing new drugs that would aid vaccines in the fight against the virus. Although there are already several potential candidates for a good inhibitor of this protein, the degree of variability of the protein itself is not taken into account.

Investigation of the P1' and P2' sites of IQF substrates and their selectivity toward 20S proteasome subunits.

High levels of expression and activity of the 20S proteasome have been linked to many types of pathologies, including neoplasia, autoimmune disorders, neurodegenerative diseases and many more. Moreover, distinguishing between 20S proteasome catalytic subunits is neglected, although it may provide further insight into the development of pathologies. Several approaches have been developed to detect 20S proteasome activity, one of which is internally quenched fluorescent (IQF) substrates, which currently suffer from low efficiency and sensitivity.

Characterization of the Ubiquitin and ISG15 Deconjugase Activity of SARS-CoV-1 and SARS-CoV-2 Papain-Like Protease.

Both severe acute respiratory syndrome coronavirus 1 and 2 (SARS-CoV-1 and SARS-CoV-2) encode a papain-like protease (PLpro), which plays a vital role in viral propagation. PLpro accomplishes this function by processing the viral polyproteins essential for viral replication and removing the small proteins, ubiquitin and ISG15 from the host's key immune signaling proteins, thereby preventing the host's innate immune response. Although PLpro from both SARS-CoV-1 and SARS-CoV-2 are structurally highly similar (83% sequence identity), they exhibit functional variability.

Design and Synthesis of Ubiquitin-Based Chemical Tools with Unnatural Amino Acids for Selective Detection of Deubiquitinases.

Several chemical approaches have been applied to develop Ub-based substrates and probes selective toward one or a narrow subset of deubiquitinases (DUBs). Since DUBs are highly specific toward ubiquitin and exhibit low activity toward shorter peptides, it is challenging to design truly selective chemical tools to investigate one DUB in biological samples. Incorporating amino acids other than canonical LRG at the P4-P2 positions in the Ub improves DUB activity and selectivity toward Ub derivatives.

Betulin and Its Derivatives Reduce Inflammation and COX-2 Activity in Macrophages.

Betulin is a heavily studied natural compound for its use as an anticancer or pro-regenerative agent. The structural similarity between betulin to steroids gives rise to the idea that the substance may as well act as an anti-inflammatory drug. This study is the first to describe the anti-inflammatory properties of betulinic acid, betulin, and its derivatives with amino acids 1,4-diaminebutane (Dab), 1,3-diaminepropane (Dap), Ornithine (Orn), and lysine (Lys) on murine macrophages from lymphoma site.

Diagnostic and therapeutic potential of protease inhibition.

Proteases are enzymes that hydrolyze peptide bonds in proteins and peptides; thus, they control virtually all biological processes. Our understanding of protease function has advanced considerably from nonselective digestive enzymes to highly specialized molecular scissors that orchestrate complex signaling networks through a limited proteolysis. The catalytic activity of proteases is tightly regulated at several levels, ranging from gene expression through trafficking and maturation to posttranslational modifications.

Evaluation of the anti-SARS-CoV-2 properties of essential oils and aromatic extracts.

Essential oils and aromatic extracts (oleoresins, absolutes, concretes, resinoids) are often used as food flavorings and constituents of fragrance compositions. The flavor and fragrance industry observed significant growth in the sales of some natural materials during the COVID-19 outbreak. Some companies worldwide are making false claims regarding the effectiveness of their essential oils or blends (or indirectly point toward this conclusion) against coronaviruses, even though the available data on the activity of plant materials against highly pathogenic human coronaviruses are very scarce.

Green-Emitting 4,5-Diaminonaphthalimides in Activity-Based Probes for the Detection of Thrombin.

The natures of electron-donating groups as well as the bridge between them determine the fate of substituted 1,8-naphthalimide molecules in the excited state. An activity-based probe constructed from a selective peptide sequence, a reactive warhead, and the brightest green-emitting fluorophore displays impressive performance for thrombin protease detection in a newly constructed series of 1,8-naphthalimides.

Parallel imaging of coagulation pathway proteases activated protein C, thrombin, and factor Xa in human plasma.

Activated protein C (APC), thrombin, and factor (f) Xa are vitamin K-dependent serine proteases that are key factors in blood coagulation. Moreover, they play important roles in inflammation, apoptosis, fibrosis, angiogenesis, and viral infections. Abnormal activity of these coagulation factors has been related to multiple conditions, such as bleeding and thrombosis, Alzheimer's disease, sepsis, multiple sclerosis, and COVID-19.

Targeting SARS-CoV-2 Proteases for COVID-19 Antiviral Development.

The emergence of severe acute respiratory syndrome (SARS-CoV-2) in 2019 marked the third occurrence of a highly pathogenic coronavirus in the human population since 2003. As the death toll surpasses 5 million globally and economic losses continue, designing drugs that could curtail infection and disease progression is critical. In the US, three highly effective Food and Drug Administration (FDA)-authorized vaccines are currently available, and Remdesivir is approved for the treatment of hospitalized patients.

Mechanistic insights into COVID-19 by global analysis of the SARS-CoV-2 3CL substrate degradome.

The main viral protease (3CL) is indispensable for SARS-CoV-2 replication. We delineate the human protein substrate landscape of 3CL by TAILS substrate-targeted N-terminomics. We identify more than 100 substrates in human lung and kidney cells supported by analyses of SARS-CoV-2-infected cells.