Coagulation modifiers targeting SARS-CoV-2 main protease Mpro for COVID-19 treatment: an in silico approach

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection depends on viral polyprotein processing, catalysed by the main proteinase (Mpro). The solution of the SARS-CoV-2 Mpro structure allowed the investigation of potential inhibitors. This work aims to provide first evidences of the applicability of commercially approved drugs to treat coronavirus disease-19 (COVID-19). We screened 4,334 compounds to found potential inhibitors of SARS-CoV-2 replication using an in silico approach. Our results evidenced the potential use of coagulation modifiers in COVID-19 treatment due to the structural similarity of SARS-CoV-2 Mpro and human coagulation factors thrombin and Factor Xa. Further in vitro and in vivo analysis are needed to corroborate these results

Kinase Inhibitor Profile For Human Nek1, Nek6, And Nek7 And Analysis Of The Structural Basis For Inhibitor Specificity.

Human Neks are a conserved protein kinase family related to cell cycle progression and cell division and are considered potential drug targets for the treatment of cancer and other pathologies. We screened the activation loop mutant kinases hNek1 and hNek2, wild-type hNek7, and five hNek6 variants in different activation/phosphorylation statesand compared them against 85 compounds using thermal shift denaturation. We identified three compounds with significant Tm shifts: JNK Inhibitor II for hNek1(Δ262-1258)-(T162A), Isogranulatimide for hNek6(S206A), andGSK-3 Inhibitor XIII for hNek7wt. Each one of these compounds was also validated by reducing the kinases activity by at least 25%. The binding sites for these compounds were identified by in silico docking at the ATP-binding site of the respective hNeks. Potential inhibitors were first screened by thermal shift assays, had their efficiency tested by a kinase assay, and were finally analyzed by molecular docking. Our findings corroborate the idea of ATP-competitive inhibition for hNek1 and hNek6 and suggest a novel non-competitive inhibition for hNek7 in regard to GSK-3 Inhibitor XIII. Our results demonstrate that our approach is useful for finding promising general and specific hNekscandidate inhibitors, which may also function as scaffolds to design more potent and selective inhibitors.201176-9

Pervasive gaps in Amazonian ecological research

Biodiversity loss is one of the main challenges of our time,1,2 and attempts to address it require a clear un derstanding of how ecological communities respond to environmental change across time and space.3,4 While the increasing availability of global databases on ecological communities has advanced our knowledge of biodiversity sensitivity to environmental changes,5–7 vast areas of the tropics remain understudied.8–11 In the American tropics, Amazonia stands out as the world’s most diverse rainforest and the primary source of Neotropical biodiversity,12 but it remains among the least known forests in America and is often underrepre sented in biodiversity databases.13–15 To worsen this situation, human-induced modifications16,17 may elim inate pieces of the Amazon’s biodiversity puzzle before we can use them to understand how ecological com munities are responding. To increase generalization and applicability of biodiversity knowledge,18,19 it is thus crucial to reduce biases in ecological research, particularly in regions projected to face the most pronounced environmental changes. We integrate ecological community metadata of 7,694 sampling sites for multiple or ganism groups in a machine learning model framework to map the research probability across the Brazilian Amazonia, while identifying the region’s vulnerability to environmental change. 15%–18% of the most ne glected areas in ecological research are expected to experience severe climate or land use changes by 2050. This means that unless we take immediate action, we will not be able to establish their current status, much less monitor how it is changing and what is being lostinfo:eu-repo/semantics/publishedVersio

Pervasive gaps in Amazonian ecological research

Biodiversity loss is one of the main challenges of our time,1,2 and attempts to address it require a clear understanding of how ecological communities respond to environmental change across time and space.3,4 While the increasing availability of global databases on ecological communities has advanced our knowledge of biodiversity sensitivity to environmental changes,5,6,7 vast areas of the tropics remain understudied.8,9,10,11 In the American tropics, Amazonia stands out as the world's most diverse rainforest and the primary source of Neotropical biodiversity,12 but it remains among the least known forests in America and is often underrepresented in biodiversity databases.13,14,15 To worsen this situation, human-induced modifications16,17 may eliminate pieces of the Amazon's biodiversity puzzle before we can use them to understand how ecological communities are responding. To increase generalization and applicability of biodiversity knowledge,18,19 it is thus crucial to reduce biases in ecological research, particularly in regions projected to face the most pronounced environmental changes. We integrate ecological community metadata of 7,694 sampling sites for multiple organism groups in a machine learning model framework to map the research probability across the Brazilian Amazonia, while identifying the region's vulnerability to environmental change. 15%–18% of the most neglected areas in ecological research are expected to experience severe climate or land use changes by 2050. This means that unless we take immediate action, we will not be able to establish their current status, much less monitor how it is changing and what is being lost

Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of the mitochondrial tryparedoxin peroxidase from Leishmania braziliensis

Submitted by Luciane Willcox ([email protected]) on 2016-09-02T17:49:47Z No. of bitstreams: 1 Cloning, expression, purification, crystallization.pdf: 350581 bytes, checksum: 0d688db5291c27edd0bfbdaf3b7372c5 (MD5)Approved for entry into archive by Luciane Willcox ([email protected]) on 2016-09-02T17:57:41Z (GMT) No. of bitstreams: 1 Cloning, expression, purification, crystallization.pdf: 350581 bytes, checksum: 0d688db5291c27edd0bfbdaf3b7372c5 (MD5)Made available in DSpace on 2016-09-02T17:57:41Z (GMT). No. of bitstreams: 1 Cloning, expression, purification, crystallization.pdf: 350581 bytes, checksum: 0d688db5291c27edd0bfbdaf3b7372c5 (MD5) Previous issue date: 2013-03-28This research was supported by grants from Fundação de Amparo a Pesquisa do Estado de São Paulo (FAPESP), Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES).Centro Nacional de Pesquisa em Energia e Materiais. Laboratório Nacional de Biociências. Campinas, SP, Brasil. / Centro Nacional de Pesquisa em Energia e Materiais. Laboratório Nacional de Luz Síncrotron. Campinas, SP, Brasil.Centro Nacional de Pesquisa em Energia e Materiais. Laboratório Nacional de Biociências. Campinas, SP, Brasil. / Centro Nacional de Pesquisa em Energia e Materiais. Laboratório Nacional de Luz Síncrotron. Campinas, SP, Brasil. / Fundação Oswaldo Cruz. Instituto Carlos Chagas. Curitiba, PR, Brasil.Centro Nacional de Pesquisa em Energia e Materiais. Laboratório Nacional de Biociências. Campinas, SP, Brasil. / Centro Nacional de Pesquisa em Energia e Materiais. Laboratório Nacional de Luz Síncrotron. Campinas, SP, Brasil.Tryparedoxin peroxidase (TXNPx) is an essential constituent of the main enzymatic scavenger system for reactive oxygen species (ROS) in trypanosomatids. Genetic studies have demonstrated the importance of this system for the development and virulence of these parasites, representing a potential target for the discovery of new trypanocidal drugs. In this work, the mitochondrial TXNPx from Leishmania braziliensis was cloned, overexpressed, purified and crystallized. The crystals diffracted to 3.3 Å resolution and belonged to space group P4(2)2(1)2, with unit-cell parameters a = b = 131.8, c = 44.4 Å. These studies will contribute to a better understanding of the molecular mechanisms involved in ROS detoxification by trypanosomatids

A Method for Purifying Native Transthyretin from Human Plasma

Submitted by Luciane Willcox ([email protected]) on 2016-08-25T15:01:25Z No. of bitstreams: 1 a-method-for-purifying-native-transthyretin-from-human-plasma-2167-0501-1000187.pdf: 828721 bytes, checksum: 9815e84d8538dc972d3fe82580b5fa2f (MD5)Approved for entry into archive by Luciane Willcox ([email protected]) on 2016-08-26T12:46:28Z (GMT) No. of bitstreams: 1 a-method-for-purifying-native-transthyretin-from-human-plasma-2167-0501-1000187.pdf: 828721 bytes, checksum: 9815e84d8538dc972d3fe82580b5fa2f (MD5)Made available in DSpace on 2016-08-26T12:46:28Z (GMT). No. of bitstreams: 1 a-method-for-purifying-native-transthyretin-from-human-plasma-2167-0501-1000187.pdf: 828721 bytes, checksum: 9815e84d8538dc972d3fe82580b5fa2f (MD5) Previous issue date: 2015-09-21Oswaldo Cruz Fundation. Institute Carlos Chagas. Laboratory of Proteomics and Proteins Engineering. Curitiba, PR, Brazil / Universidade Federal do Paraná. Curitiba, PR, Brazil.Oswaldo Cruz Fundation. Institute Carlos Chagas. Laboratory of Proteomics and Proteins Engineering. Curitiba, PR, Brazil.Oswaldo Cruz Fundation. Institute Carlos Chagas. Laboratory of Proteomics and Proteins Engineering. Curitiba, PR, Brazil.Oswaldo Cruz Fundation. Institute Carlos Chagas. Laboratory of Proteomics and Proteins Engineering. Curitiba, PR, Brazil.Transthyretin is a homotetrameric thyroid-hormone-transporting protein that binds to the retinol binding protein thus being involved in metabolism, growth, fertility, homeostasis of the cardiovascular and central nervous system, cell differentiation, reproduction, development and maintenance the cognitive processes during aging. Currently, there are several methodologies for natively purifying TTR from plasma, serum, tears, and amyloid fibrils; however, these procedures are laborious. Herein, a low-cost and simple protocol to purify TTR from human plasma is described. It involves the separation of plasma proteins by size exclusion and DEAE chromatography. The homogeneity was assessed by SDS-PAGE and by tandem mass spectrometry using an Orbitrap-XL (Thermo, San Jose-CA)

Purification, crystallization and preliminary X-ray diffraction analysis of a class P-III metalloproteinase (BmMP-III) from the venom of Bothrops moojeni

Snake-venom metalloproteinases (SVMPs) comprise a family of haemostatically active toxins which can cause haemorrhage, coagulopathy, inhibition of platelet aggregation and inflammatory response. These effects are attributed to the proteolytic action of SVMPs on extracellular matrix components, plasma proteins and cell-surface proteins. SVMPs are classified into four classes (P-I to P-IV) based on their domain structures. In order to understand the multiple roles played by the domains of P-III SVMPs, a P-III SVMP (BmMP-III) from the venom of Bothrops moojeni was purified, characterized and crystallized. The crystals belonged to space group I4(1)22, with unit-cell parameters a = b = 108.16, c = 196.09 angstrom. Initially, flash-cooled crystals diffracted poorly to a resolution of about 10 angstrom. However, a significant improvement in the diffraction resolution was observed upon annealing and a complete data set was collected to 3.3 angstrom resolution. The asymmetric unit contained one molecule and the structure was determined and partially refined to an R factor of 34%. Structural comparisons indicated that the cysteine-rich domain can adopt different conformations in relation to the catalytic domain, which may modulate the enzyme activity.Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES