1. Identification of triazolopyrimidinyl scaffold SARS-CoV-2 papain-like protease (PLpro) inhibitorSebastjan Kralj, Marko Jukič, Miha Bahun, Luka Kranjc, Anja Kolarič, Milan Hodošček, Nataša Poklar Ulrih, Urban Bren, 2024, original scientific article Abstract: The global impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its companion disease, COVID-19, has reminded us of the importance of basic coronaviral research. In this study, a comprehensive approach using molecular docking, in vitro assays, and molecular dynamics simulations was applied to identify potential inhibitors for SARS-CoV-2 papain-like protease (PLpro), a key and underexplored viral enzyme target. A focused protease inhibitor library was initially created and molecular docking was performed using CmDock software (v0.2.0), resulting in the selection of hit compounds for in vitro testing on the isolated enzyme. Among them, compound 372 exhibited promising inhibitory properties against PLpro, with an IC50 value of 82 ± 34 μM. The compound also displayed a new triazolopyrimidinyl scaffold not yet represented within protease inhibitors. Molecular dynamics simulations demonstrated the favorable binding properties of compound 372. Structural analysis highlighted its key interactions with PLpro, and we stress its potential for further optimization. Moreover, besides compound 372 as a candidate for PLpro inhibitor development, this study elaborates on the PLpro binding site dynamics and provides a valuable contribution for further efforts in pan-coronaviral PLpro inhibitor development.
Keywords: drug design, protease inhibitor, SARS-CoV-2, papain-like protease, PLpro, antiviral design, in silico drug design, CADD, virtual screening, HTVS, structure-based design
Published in DiRROS: 07.08.2024; Views: 88; Downloads: 96
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2. Proteases and cytokines as mediators of interactions between cancer and stromal cells in tumoursBarbara Breznik, Helena Motaln, Tamara Lah Turnšek, 2017, review article Abstract: Proteolytic enzymes are highly relevant in different processes of cancer progression. Their interplay with other signalling molecules such as cytokines represents important regulation of multicellular cross-talk. In this review, we discuss protease regulation mechanisms of cytokine signalling in various types of cancer. Additionally, we highlight the reverse whereby cytokines have an impact on protease expression in an autocrine and paracrine manner, representing complex feedback mechanisms among multiple members of these two protein families. The relevance of the protease-cytokine axis is illustrated in glioblastoma, where interactions between normal mesenchymal stem cells and cancer cells play an important role in this very malignant form of brain cancer.
Keywords: cellular cross-talk, glioblastoma, invasion, mesenchymal stem cells, protease-cytokine signalling
Published in DiRROS: 06.08.2024; Views: 95; Downloads: 62
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3. Gene expression profiling of recombinant protein producing E. coli at suboptimal growth temperatureMitja Mahnič, Špela Baebler, Andrej Blejec, Špela Jalen, Kristina Gruden, Viktor Menart, Simona Jevševar, 2012, original scientific article Abstract: Recent studies have revealed that at lower cultivation temperatures (25 °C) much higher percentage of correctly folded recombinant hG-CSF protein can be extracted from inclusion bodies. Hence, the goal of our research was to investigate mechanisms determining characteristics of non-classical inclusion bodies production using gene expression profiling, focusing on proteases and chaperones gene expression. Statistical analysis of microarray data showed prominent changes in energy metabolism, in metabolism of amino acids and nucleotides, as well as in biosynthesis of cofactors and secondary metabolites if the culture was grown below its optimal temperature. Moreover, 24 differentially expressed up to now known genes classified among proteases, chaperones and other heat or stress related genes. Among chaperones UspE and among proteases YaeL and YeaZ might play an important role in accumulation of correctly folded recombinant proteins. Membrane localized protease yaeL gene was found to have higher activity at 25 °C and is thus potentially functionally related to the more efficient recombinant protein production at lower temperatures. The results of this study represent advance in the understanding of recombinant protein production in E. coli. Genes potentially influencing production of recombinant protein at lower growth temperature represent basis for further research towards improvement of E. coli production strains as well as fermentation process.
Keywords: recombinant protein production, non-classical inclusion bodies, expression microarrays, YaeL protease, GroEL chaperone
Published in DiRROS: 05.08.2024; Views: 221; Downloads: 206
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4. Stress-tolerant yeasts : opportunistic pathogenicity versus biocontrol potentialJanja Zajc, Cene Gostinčar, Anja Černoša, Nina Gunde-Cimerman, 2019, original scientific article Abstract: Stress-tolerant fungi that can thrive under various environmental extremes are highly desirable for their application to biological control, as an alternative to chemicals for pest management. However, in fungi, the mechanisms of stress tolerance might also have roles in mammal opportunism. We tested five species with high biocontrol potential in agriculture (Aureobasidium pullulans, Debayomyces hansenii, Meyerozyma guilliermondii, Metschnikowia fructicola, Rhodotorula mucilaginosa) and two species recognized as emerging opportunistic human pathogens (Exophiala dermatitidis, Aureobasidium melanogenum) for growth under oligotrophic conditions and at 37 °C, and for tolerance to oxidative stress, formation of biofilms, production of hydrolytic enzymes and siderophores, and use of hydrocarbons as sole carbon source. The results show large overlap between traits desirable for biocontrol and traits linked to opportunism (growth under oligotrophic conditions, production of siderophores, high oxidative stress tolerance, and specific enzyme activities). Based on existing knowledge and these data, we suggest that oligotrophism and thermotolerance together with siderophore production at 37 °C, urease activity, melanization, and biofilm production are the main traits that increase the potential for fungi to cause opportunistic infections in mammals. These traits should be carefully considered when assessing safety of potential biocontrol agents.
Keywords: opportunistic pathogen, biocontrol agent, virulence, stress tolerance, secretome, CAZy, protease, thermotolerance, oligotrophism, melanin, siderophore, biofilm
Published in DiRROS: 23.07.2024; Views: 128; Downloads: 139
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5. Proteolytically activated CRAC effectors through designed intramolecular inhibitionVid Jazbec, Roman Jerala, Mojca Benčina, 2022, original scientific article Keywords: STIM1, Orai, TEV protease, PPV protease, calcium signaling, coiled-coil peptides
Published in DiRROS: 20.07.2022; Views: 877; Downloads: 528
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