Some medically important viruses-including retroviruses, flaviviruses, coronaviruses, and herpesviruses-code for a protease, which is indispensable for viral maturation and pathogenesis. Viral protease inhibitors have become an important class of antiviral drugs. Development of the first-in-class viral protease inhibitor saquinavir, which targets HIV protease, started a new era in the treatment of chronic viral diseases. Combining several drugs that target different steps of the viral life cycle enables use of lower doses of individual drugs (and thereby reduction of potential side effects, which frequently occur during long term therapy) and reduces drug-resistance development. Currently, several HIV and HCV protease inhibitors are routinely used in clinical practice. In addition, a drug including an inhibitor of SARS-CoV-2 main protease, nirmatrelvir (co-administered with a pharmacokinetic booster ritonavir as Paxlovid®), was recently authorized for emergency use. This review summarizes the basic features of the proteases of human immunodeficiency virus (HIV), hepatitis C virus (HCV), and SARS-CoV-2 and discusses the properties of their inhibitors in clinical use, as well as development of compounds in the pipeline.

• MeSH
• antivirové látky farmakologie   terapeutické užití   MeSH
• COVID-19 * MeSH
• HIV infekce * farmakoterapie   MeSH
• lidé MeSH
• SARS-CoV-2 MeSH
• virové proteasy MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

The worldwide COVID-19 pandemic caused by the coronavirus SARS-CoV-2 urgently demands novel direct antiviral treatments. The main protease (Mpro) and papain-like protease (PLpro) are attractive drug targets among coronaviruses due to their essential role in processing the polyproteins translated from the viral RNA. In this study, we virtually screened 688 naphthoquinoidal compounds and derivatives against Mpro of SARS-CoV-2. Twenty-four derivatives were selected and evaluated in biochemical assays against Mpro using a novel fluorogenic substrate. In parallel, these compounds were also assayed with SARS-CoV-2 PLpro. Four compounds inhibited Mpro with half-maximal inhibitory concentration (IC50) values between 0.41 μM and 9.0 μM. In addition, three compounds inhibited PLpro with IC50 ranging from 1.9 μM to 3.3 μM. To verify the specificity of Mpro and PLpro inhibitors, our experiments included an assessment of common causes of false positives such as aggregation, high compound fluorescence, and inhibition by enzyme oxidation. Altogether, we confirmed novel classes of specific Mpro and PLpro inhibitors. Molecular dynamics simulations suggest stable binding modes for Mpro inhibitors with frequent interactions with residues in the S1 and S2 pockets of the active site. For two PLpro inhibitors, interactions occur in the S3 and S4 pockets. In summary, our structure-based computational and biochemical approach identified novel naphthoquinonal scaffolds that can be further explored as SARS-CoV-2 antivirals.

• MeSH
• antivirové látky * farmakologie   chemie   MeSH
• COVID-19 MeSH
• inhibitory proteas * farmakologie   chemie   MeSH
• koronavirové proteasy 3C * antagonisté a inhibitory   MeSH
• koronavirové proteasy podobné papainu * antagonisté a inhibitory   MeSH
• lidé MeSH
• naftochinony * chemie   farmakologie   MeSH
• papain MeSH
• SARS-CoV-2 * účinky léků   enzymologie   MeSH
• simulace molekulového dockingu MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH

Specific salts effect is intensively studied from the prospective of modification of different physico-chemical properties of biomacromolecules. Limited knowledge of the specific salts effect on enzymes led us to address the influence of five sodium anions: sulfate, phosphate, chloride, bromide, and perchlorate, on catalytic and conformational properties of human rhinovirus-14 (HRV) 3C protease. The enzyme conformation was monitored by circular dichroism spectrum (CD) and by tyrosines fluorescence. Stability and flexibility of the enzyme have been analyzed by CD in the far-UV region, differential scanning calorimetry and molecular dynamics simulations, respectively. We showed significant influence of the anions on the enzyme properties in accordance with the Hofmeister effect. The HRV 3C protease in the presence of kosmotropic anions, in contrast with chaotropic anions, exhibits increased stability, rigidity. Correlations of stabilization effect of anions on the enzyme with their charge density and the rate constant of the enzyme with the viscosity B-coefficients of anions suggest direct interaction of the anions with HRV 3C protease. The role of stabilization and decreased fluctuation of the polypeptide chain of HRV 3C protease on its activation in the presence of kosmotropic anions is discussed within the frame of the macromolecular rate theory.

• MeSH
• anionty chemie   MeSH
• lidé MeSH
• prospektivní studie MeSH
• proteasy * MeSH
• soli * chemie   MeSH
• srdeční frekvence MeSH
• virové proteasy 3C MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Drug repurposing requires a limited resource, cost-effective and faster method to combat severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Therefore, this in silico studies attempts to identify the drug-likeness properties of ravidasvir, an II/III phase clinical trial chronic hepatitis C drug against 3-Chymotrypsin-like protease (3CLpro) of SARS-CoV-2 to combat the ongoing coronavirus disease 2019 (COVID-19) pandemic. This protease is predominantly involved in virus replication cycle; hence it is considered as a potent drug target. The molecular docking results showed that ravidasvir was found to be potent inhibitors of 3CLpro with scoring function based binding energy is -26.7 kJ/mol. Further dynamic behaviour of apo form and complex form of ravidasvir with 3CLpro were studied using molecular dynamics (MD) simulations over 500 ns each, total 2 μs time scale. The motion of the protein was studied using principal component analysis of the MD simulation trajectories. The binding free energy calculated using MM/PBSA method from the MD simulation trajectory was -190.3 ± 70.2 kJ/mol and -106.0 ± 26.7 kJ/mol for GROMOS96 54A7 and AMBER99SB-ILDN force field, respectively. This in silico studies suggesting ravidasvir might be a potential lead molecule against SARS-CoV-2 for further optimization and drug development to combat the life-threatening COVID-19 pandemic.Communicated by Ramaswamy H. Sarma.

• MeSH
• adipáty MeSH
• benzimidazoly MeSH
• COVID-19 * MeSH
• cysteinové endopeptidasy chemie   MeSH
• farmakoterapie COVID-19 MeSH
• inhibitory proteas chemie   MeSH
• koronavirové proteasy 3C MeSH
• lidé MeSH
• pandemie MeSH
• SARS-CoV-2 * MeSH
• simulace molekulární dynamiky MeSH
• simulace molekulového dockingu MeSH
• sukcináty MeSH
• valin analogy a deriváty   MeSH
• virové nestrukturální proteiny chemie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The acute respiratory syndrome caused by the SARS-CoV-2, known as COVID-19, has been ruthlessly tormenting the world population for more than six months. However, so far no effective drug or vaccine against this plague have emerged yet, despite the huge effort in course by researchers and pharmaceutical companies worldwide. Willing to contribute with this fight to defeat COVID-19, we performed a virtual screening study on a library containing Food and Drug Administration (FDA) approved drugs, in a search for molecules capable of hitting three main molecular targets of SARS-CoV-2 currently available in the Protein Data Bank (PDB). Our results were refined with further molecular dynamics (MD) simulations and MM-PBSA calculations and pointed to 7 multi-target hits which we propose here for experimental evaluation and repurposing as potential drugs against COVID-19. Additional rounds of docking, MD simulations and MM-PBSA calculations with remdesivir suggested that this compound can also work as a multi-target drug against SARS-CoV-2.Communicated by Ramaswamy H. Sarma.

• MeSH
• COVID-19 * MeSH
• cysteinové endopeptidasy MeSH
• farmakoterapie COVID-19 MeSH
• inhibitory proteas MeSH
• koronavirové proteasy 3C MeSH
• léčivé přípravky MeSH
• lidé MeSH
• SARS-CoV-2 * MeSH
• simulace molekulární dynamiky MeSH
• simulace molekulového dockingu MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Viral proteases are indispensable for successful virion maturation, thus making them a prominent drug target. Their enzyme activity is tightly spatiotemporally regulated by expression in the precursor form with little or no activity, followed by activation via autoprocessing. These cleavage events are frequently triggered upon transportation to a specific compartment inside the host cell. Typically, precursor oligomerization or the presence of a co-factor is needed for activation. A detailed understanding of these mechanisms will allow ligands with non-canonical mechanisms of action to be designed, which would specifically modulate the initial irreversible steps of viral protease autoactivation. Binding sites exclusive to the precursor, including binding sites beyond the protease domain, can be exploited. Both inhibition and up-regulation of the proteolytic activity of viral proteases can be detrimental for the virus. All these possibilities are discussed using examples of medically relevant viruses including herpesviruses, adenoviruses, retroviruses, picornaviruses, caliciviruses, togaviruses, flaviviruses, and coronaviruses.

• MeSH
• antivirové látky farmakologie   MeSH
• Flavivirus účinky léků   metabolismus   MeSH
• Herpesviridae účinky léků   metabolismus   MeSH
• HIV-1 účinky léků   MeSH
• inhibitory virových proteáz farmakologie   MeSH
• lidé MeSH
• lidské adenoviry účinky léků   metabolismus   MeSH
• SARS-CoV-2 účinky léků   metabolismus   MeSH
• virové nemoci farmakoterapie   MeSH
• virové proteasy biosyntéza   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

Spanish flu, polio epidemics, and the ongoing COVID-19 pandemic are the most profound examples of severe widespread diseases caused by RNA viruses. The coronavirus pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) demands affordable and reliable assays for testing antivirals. To test inhibitors of viral proteases, we have developed an inexpensive high-throughput assay based on fluorescent energy transfer (FRET). We assayed an array of inhibitors for papain-like protease from SARS-CoV-2 and validated it on protease from the tick-borne encephalitis virus to emphasize its versatility. The reaction progress is monitored as loss of FRET signal of the substrate. This robust and reproducible assay can be used for testing the inhibitors in 96- or 384-well plates.

• MeSH
• antivirové látky farmakologie   MeSH
• COVID-19 MeSH
• farmakoterapie COVID-19 MeSH
• fluorescenční barviva chemie   MeSH
• inhibitory proteas farmakologie   MeSH
• koronavirové proteasy podobné papainu antagonisté a inhibitory   chemie   genetika   metabolismus   MeSH
• lidé MeSH
• preklinické hodnocení léčiv MeSH
• rezonanční přenos fluorescenční energie metody   MeSH
• RNA-helikasy antagonisté a inhibitory   chemie   genetika   metabolismus   MeSH
• RNA-viry enzymologie   MeSH
• rychlé screeningové testy metody   MeSH
• SARS-CoV-2 enzymologie   MeSH
• serinové endopeptidasy chemie   genetika   metabolismus   MeSH
• virové nestrukturální proteiny antagonisté a inhibitory   chemie   genetika   metabolismus   MeSH
• viry klíšťové encefalitidy enzymologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Cryptococcosis is an invasive infection that accounts for 15% of AIDS-related fatalities. Still, treating cryptococcosis remains a significant challenge due to the poor availability of effective antifungal therapies and emergence of drug resistance. Interestingly, protease inhibitor components of antiretroviral therapy regimens have shown some clinical benefits in these opportunistic infections. We investigated Major aspartyl peptidase 1 (May1), a secreted Cryptococcus neoformans protease, as a possible target for the development of drugs that act against both fungal and retroviral aspartyl proteases. Here, we describe the biochemical characterization of May1, present its high-resolution X-ray structure, and provide its substrate specificity analysis. Through combinatorial screening of 11,520 compounds, we identified a potent inhibitor of May1 and HIV protease. This dual-specificity inhibitor exhibits antifungal activity in yeast culture, low cytotoxicity, and low off-target activity against host proteases and could thus serve as a lead compound for further development of May1 and HIV protease inhibitors.

• MeSH
• antifungální látky chemie   metabolismus   farmakologie   MeSH
• aspartátové proteasy antagonisté a inhibitory   genetika   metabolismus   MeSH
• Cryptococcus neoformans enzymologie   MeSH
• fungální proteiny antagonisté a inhibitory   genetika   metabolismus   MeSH
• HIV-proteasa chemie   metabolismus   MeSH
• HIV enzymologie   MeSH
• houby účinky léků   MeSH
• katalytická doména MeSH
• krystalografie rentgenová MeSH
• preklinické hodnocení léčiv MeSH
• rekombinantní proteiny biosyntéza   chemie   izolace a purifikace   MeSH
• simulace molekulární dynamiky MeSH
• substrátová specifita MeSH
• vazebná místa MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH

• Klíčová slova
• nezralé částice, inhibitory schopnosti tvořit částice,
• MeSH
• fluorescenční mikroskopie MeSH
• genové produkty gag - virus lidské imunodeficience MeSH
• HIV infekce epidemiologie   etiologie   terapie   MeSH
• HIV-proteasa MeSH
• HIV genetika   patogenita   růst a vývoj   účinky léků   MeSH
• látky proti HIV * farmakologie   MeSH
• léková rezistence MeSH
• lidé MeSH
• mutantní proteiny MeSH
• objevování léků * metody   MeSH
• techniky in vitro MeSH
• testy rezistence k nukleáze MeSH
• vysoce aktivní antiretrovirová terapie ekonomika   škodlivé účinky   MeSH
• Check Tag
• lidé MeSH

High-pressure methods have become an interesting tool of investigation of structural stability of proteins. They are used to study protein unfolding, but dissociation of oligomeric proteins can be addressed this way, too. HIV-1 protease, although an interesting object of biophysical experiments, has not been studied at high pressure yet. In this study HIV-1 protease is investigated by high pressure (up to 600 MPa) fluorescence spectroscopy of either the inherent tryptophan residues or external 8-anilino-1-naphtalenesulfonic acid at 25°C. A fast concentration-dependent structural transition is detected that corresponds to the dimer-monomer equilibrium. This transition is followed by a slow concentration independent transition that can be assigned to the monomer unfolding. In the presence of a tight-binding inhibitor none of these transitions are observed, which confirms the stabilizing effect of inhibitor. High-pressure enzyme kinetics (up to 350 MPa) also reveals the stabilizing effect of substrate. Unfolding of the protease can thus proceed only from the monomeric state after dimer dissociation and is unfavourable at atmospheric pressure. Dimer-destabilizing effect of high pressure is caused by negative volume change of dimer dissociation of -32.5 mL/mol. It helps us to determine the atmospheric pressure dimerization constant of 0.92 μM. High-pressure methods thus enable the investigation of structural phenomena that are difficult or impossible to measure at atmospheric pressure.

• MeSH
• anilin-naftalen sulfonáty metabolismus   MeSH
• atmosférický tlak MeSH
• darunavir metabolismus   MeSH
• dimerizace MeSH
• fluorescenční spektrometrie MeSH
• HIV-proteasa chemie   metabolismus   MeSH
• inhibitory HIV-proteasy metabolismus   MeSH
• kinetika MeSH
• konformace proteinů MeSH
• lidé MeSH
• molekulární modely MeSH
• multimerizace proteinu MeSH
• sbalování proteinů * MeSH
• stabilita proteinů účinky léků   MeSH
• termodynamika MeSH
• tryptofan metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH