35 research outputs found
Binding adaptation of GS 441524 diversifies macro domains and downregulate SARS CoV 2 de MARylation capacity
Viral infection in cells triggers a cascade of molecular defense mechanisms to maintain host cell homoeostasis. One of these mechanisms is ADP ribosylation, a fundamental post translational modification PTM characterized by the addition of ADP ribose ADPr on substrates. Poly ADP ribose polymerases PARPs are implicated in this process and they perform ADP ribosylation on host and pathogen proteins. Some viral families contain structural motifs that can reverse this PTM. These motifs known as macro domains MDs are evolutionarily conserved protein domains found in all kingdoms of life. They are divided in different classes with the viral belonging to Macro D type class because of their properties to recognize and revert the ADP ribosylation. Viral MDs are potential pharmaceutical targets, capable to counteract host immune response. Sequence and structural homology between viral and human MDs are an impediment for the development of new active compounds against their function. Remdesivir, is a drug administrated in viral infections inhibiting viral replication through RNA dependent RNA polymerase RdRp . Herein, GS 441524, the active metabolite of the remdesivir, is tested as a hydrolase inhibitor for several viral MDs and for its binding to human homologs found in PARPs. This study presents biochemical and biophysical studies, which indicate that GS 441524 selectively modifies SARS CoV 2 MD de MARylation activity, while it does not interact with hPARP14 MD2 and hPARP15 MD2. The structural investigation of MD GS 441524 complexes, using solution NMR and X ray crystallography, discloses the impact of certain amino acids in ADPr binding cavity suggesting that F360 and its adjacent residues tune the selective binding of the inhibitor to SARS CoV 2 M
Comprehensive Fragment Screening of the SARS-CoV-2 Proteome Explores Novel Chemical Space for Drug Development
12 pags., 4 figs., 3 tabs.SARS-CoV-2 (SCoV2) and its variants of concern pose serious challenges to the public health. The variants increased challenges to vaccines, thus necessitating for development of new intervention strategies including anti-virals. Within the international Covid19-NMR consortium, we have identified binders targeting the RNA genome of SCoV2. We established protocols for the production and NMR characterization of more than 80â% of all SCoV2 proteins. Here, we performed an NMR screening using a fragment library for binding to 25 SCoV2 proteins and identified hits also against previously unexplored SCoV2 proteins. Computational mapping was used to predict binding sites and identify functional moieties (chemotypes) of the ligands occupying these pockets. Striking consensus was observed between NMR-detected binding sites of the main protease and the computational procedure. Our investigation provides novel structural and chemical space for structure-based drug design against the SCoV2 proteome.Work at BMRZ is supported by the state of Hesse. Work in Covid19-NMR
was supported by the Goethe Corona Funds, by the IWBEFRE-program 20007375 of state of Hesse, the DFG
through CRC902: âMolecular Principles of RNA-based regulation.â and through infrastructure funds (project
numbers: 277478796, 277479031, 392682309, 452632086, 70653611) and by European Unionâs Horizon 2020 research and innovation program iNEXT-discovery under grant agreement No 871037. BY-COVID receives funding from the European Unionâs Horizon Europe Research and Innovation Programme under grant agreement number 101046203. âINSPIREDâ (MIS 5002550) project, implemented under the Action âReinforcement of the Research and Innovation Infrastructure,â funded by the Operational
Program âCompetitiveness, Entrepreneurship and Innovationâ (NSRF 2014â2020) and co-financed by Greece and the EU (European Regional Development Fund) and the FP7 REGPOT CT-2011-285950ââSEE-DRUGâ project (purchase of UPATâs 700 MHz NMR equipment). The support of the CERM/CIRMMP center of Instruct-ERIC is gratefully acknowledged. This work has been funded in part by a grant of the Italian Ministry of University and Research (FISR2020IP_02112, ID-COVID) and by Fondazione CR
Firenze. A.S. is supported by the Deutsche Forschungsgemeinschaft [SFB902/B16, SCHL2062/2-1] and the Johanna Quandt Young Academy at Goethe [2019/AS01]. M.H. and C.F. thank SFB902 and the Stiftung Polytechnische Gesellschaft for the Scholarship. L.L. work was supported by the French National Research Agency (ANR, NMR-SCoV2-ORF8), the Fondation de la Recherche MĂ©dicale (FRM, NMR-SCoV2-ORF8), FINOVI and the IR-RMN-THC Fr3050 CNRS. Work at UConn Health was supported by grants from the US National Institutes of Health (R01 GM135592 to B.H., P41 GM111135 and R01 GM123249 to J.C.H.) and the US National Science Foundation (DBI 2030601 to J.C.H.). Latvian Council of Science Grant No. VPP-COVID-2020/1-0014. National Science Foundation EAGER MCB-2031269. This work was supported by the grant Krebsliga KFS-4903-08-2019 and SNF-311030_192646 to J.O. P.G. (ITMP) The EOSC Future project is co-funded by the European Union Horizon Programme call INFRAEOSC-03-2020âGrant Agreement
Number 101017536. Open Access funding enabled and organized by Projekt DEALPeer reviewe
Large-Scale Recombinant Production of the SARS-CoV-2 Proteome for High-Throughput and Structural Biology Applications
The highly infectious disease COVID-19 caused by the Betacoronavirus SARS-CoV-2 poses a severe threat to humanity and demands the redirection of scientific efforts and criteria to organized research projects. The international COVID19-NMR consortium seeks to provide such new approaches by gathering scientific expertise worldwide. In particular, making available viral proteins and RNAs will pave the way to understanding the SARS-CoV-2 molecular components in detail. The research in COVID19-NMR and the resources provided through the consortium are fully disclosed to accelerate access and exploitation. NMR investigations of the viral molecular components are designated to provide the essential basis for further work, including macromolecular interaction studies and high-throughput drug screening. Here, we present the extensive catalog of a holistic SARS-CoV-2 protein preparation approach based on the consortiumâs collective efforts. We provide protocols for the large-scale production of more than 80% of all SARS-CoV-2 proteins or essential parts of them. Several of the proteins were produced in more than one laboratory, demonstrating the high interoperability between NMR groups worldwide. For the majority of proteins, we can produce isotope-labeled samples of HSQC-grade. Together with several NMR chemical shift assignments made publicly available on covid19-nmr.com, we here provide highly valuable resources for the production of SARS-CoV-2 proteins in isotope-labeled form
results of the German ConnÂŽs Registry
PrimÀrer Hyperaldosteronismus (PA) hat nachteilige Auswirkungen auf die
Nierenfunktion, die aufgrund ihrer AusprÀgung nicht allein durch die erhöhten
Blutdruckwerte erklÀrbar sind. Bisher ist die Datenlage hinsichtlich primÀrem
Hyperaldosteronismus und Nierenfunktion bzw. deren Entwicklung unter
verschiedenen Therapiearten jedoch noch dĂŒrftig. Diese prospektive,
multizentrische Studie des Deutschen Conn-Registers umfasste 29 Patienten mit
neu diagnostiziertem PA vor und ein Jahr nach Beginn der Behandlung, sowie
eine zweite Kohorte von 119 Patienten, die 5,3 und 6,8 Jahre nach Beginn der
Behandlung untersucht wurden. Es wurden die glomerulÀre Filtrationsrate (GFR),
der Albumin-Kreatinin-Quotient im Spontanurin und andere biochemische und
klinische Parameter erhoben. In einer gröĂeren Querschnittsstudie wurde die
Nierenfunktion abhÀngig von der Art der Behandlung ausgewertet (Adrenalektomie
(ADX, n = 86), Spironolacton (n = 65) und Eplerenon (n = 18)). Die GFR und der
Albumin-Kreatinin-Quotient sanken bei den neu diagnostizierten Patienten nach
Beginn der Behandlung deutlich ab. In Kohorte 2 verblieben die GFR und der
Albumin-Kreatinin-Quotient wÀhrend des Beobachtungszeitraumes stabil, die
Blutdruckwerte waren gut eingestellt. In der Querschnittsanalyse der
Gesamtkohorte wurden keine Unterschiede bzgl. GFR und Albumin-Kreatinin-
Quotient zwischen den PA Patienten mit unterschiedlichen Behandlungsschemata
gesehen. Allerdings zeigten sich unter Eplerenonbehandlung niedrigere
Kaliumwerte und eine höhere Anzahl benötigter Antihypertensiva.
Zusammenfassend lÀsst sich sagen, dass die beobachtete BeeintrÀchtigung der
Nierenfunktion und die Albuminurie der Patienten durch Beginn einer
spezifischen Behandlung reversibel waren. Die konservative Therapie mit
Spironolacton oder Eplerenon scheint bei ausreichend hoher Dosierung ebenso
effektiv zu sein wie die DurchfĂŒhrung einer Adrenalektomie was die
Blutdruckeinstellung sowie die Verbesserung der Nierenfunktion betrifft.Primary aldosteronism (PA) has deleterious effects on kidney function
independent of blood pressure levels. Up to now, data on effectiveness of
different PA therapies regarding renal function are scarce. This prospective
multi-center study of the German Connâs Registry included 29 patients with
newly diagnosed PA evaluated before and 1 year after treatment initiation, and
a second cohort including 119 patients who were evaluated 5.3 and 6.8 years
after treatment initiation. Glomerular filtration rate (GFR), spot urine
albumin excretion/urinary creatinine (UAE/Ucrea) ratio and other biochemical
and clinical parameters were measured. In a larger cross-sectional cohort,
renal function was evaluated depending on the type of treatment (adrenalectomy
(ADX; n=86), spironolactone (n=65) or eplerenone (n=18)). GFR and UAE/Ucrea
ratio significantly decreased in newly diagnosed PA patients after treatment
initiation. In the second cohort, GFR and UAE/Ucrea ratio did not change
during study period, and blood pressure was well controlled. In the larger
cross-sectional cohort, no differences were seen in GFR and UAE/Ucrea ratio
between PA patients on different treatment regimens. However, eplerenone
treatment showed lower potassium levels and higher number of required
antihypertensive medications. In conclusion, renal dysfunction with elevated
albuminuria was seen in PA patients and was reversible after treatment
initiation. Medical therapies with spironolactone or eplerenone seem to be as
effective as ADX regarding renal function and blood pressure; however,
sufficient daily doses need to be given
NMR study of macro domains (MDs) from betacoronavirus: backbone resonance assignments of SARSâCoV and MERSâCoV MDs in the free and the ADPr-bound state
Study of the soils and survey of the microrelief in the western part of the Forest of Soignes (Belgium) : with special attention to the uprooting of trees and the suitability for tree plantations /
Proefschrift voorgelegd ter verkrijging van van het diploma van licentiaat in de bodemkund