Ligand-Induced Stabilization of the Native Human Superoxide Dismutase 1

Santur, Karoline; Reinartz, Elke; Lien, Yi; Tusche, Markus; Altendorf, Tim; Sevenich, Marc; Tamgüney, Gültekin; Mohrlüder, Jeannine; Willbold, Dieter

doi:10.1021/acschemneuro.1c00253

Cited by 10 publications

(19 citation statements)

References 35 publications

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“…Variable DNA sequences resulting from next generation sequencing (NGS) were transcribed into peptide sequences, as described previously [ 36 ]. The transcribed sequences were filtered based on their relative frequency increase within the TS (library < TS1 < TS2 < TS3), the relative frequency of a sequence observed in one TS compared to its abundancy within the respective DC (TS2 > DC2; TS3 > DC3) and the relative frequency of one sequence observed in one TS round compared to its abundancy within the respective ES round (TS1 > ES1; TS2 > ES2; TS3 > ES3).…”

Section: Methodsmentioning

confidence: 99%

Phage Display-Derived Compounds Displace hACE2 from Its Complex with SARS-CoV-2 Spike Protein

et al. 2022

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Severe respiratory syndrome coronavirus-2 (SARS-CoV-2) is a highly contagious beta-class coronavirus. Although vaccinations have shown high efficacy, the emergence of novel variants of concern (VOCs) has already exhibited traits of immune evasion. Thus, the development of tailored antiviral medications for patients with incomplete, inefficient, or non-existent immunization, is essential. The attachment of viral surface proteins to the cell surface is the first crucial step in the viral replication cycle, which for SARS-CoV-2 is mediated by the high affinity interaction of the viral trimeric spike with the host cell surface-located human angiotensin converting enzyme-2 (hACE2). Here, we used a novel and efficient next generation sequencing (NGS) supported phage display strategy for the selection of a set of SARS-CoV-2 receptor binding domain (RBD)-targeting peptide ligands that bind to the target protein with low µM to nM dissociation constants. Compound CVRBDL-3 inhibits the SARS-CoV-2 spike protein association to hACE2 in a concentration-dependent manner for pre- as well as post-complex formation conditions. Further rational optimization yielded a CVRBDL-3 based divalent compound, which demonstrated inhibitory efficacy with an IC50 value of 47 nM. The obtained compounds were not only efficient for the different spike constructs from the originally isolated “wt” SARS-CoV-2, but also for B.1.1.7 mutant trimeric spike protein. Our work demonstrates that phage display-derived peptide ligands are potential fusion inhibitors of viral cell entry. Moreover, we show that rational optimization of a combination of peptide sequences is a potential strategy in the further development of therapeutics for the treatment of acute COVID-19.

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Section: Methodsmentioning

confidence: 99%

Phage Display-Derived Compounds Displace hACE2 from Its Complex with SARS-CoV-2 Spike Protein

et al. 2022

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“…The selected phages from the phage library presenting 16mer peptides on their surface were analyzed via Next Generation Sequencing (NGS). The sequencing data were evaluated via TSAT, evaluation software developed by our work group [ 42 ] and the clustering tool Hammock [ 43 ]. There was no general sequence alignment of all selected sequences possible.…”

Section: Resultsmentioning

confidence: 99%

“…The single-stranded phage DNA was prepared for analysis by next generation sequencing (NGS) as described previously [ 42 ].…”

Section: Methodsmentioning

confidence: 99%

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Inhibition of Polyglutamine Misfolding with D-Enantiomeric Peptides Identified by Mirror Image Phage Display Selection

2022

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Nine heritable diseases are known that are caused by unphysiologically elongated polyglutamine tracts in human proteins leading to misfolding, aggregation and neurodegeneration. Current therapeutic strategies include efforts to inhibit the expression of the respective gene coding for the polyglutamine-containing proteins. There are, however, concerns that this may interfere with the physiological function of the respective protein. We aim to stabilize the protein’s native conformation by D-enantiomeric peptide ligands to prevent misfolding and aggregation, shift the equilibrium between aggregates and monomers towards monomers and dissolve already existing aggregates into non-toxic and functional monomers. Here, we performed a mirror image phage display selection on the polyglutamine containing a fragment of the androgen receptor. An elongated polyglutamine tract in the androgen receptor causes spinal and bulbar muscular atrophy (SBMA). The selected D-enantiomeric peptides were tested for their ability to inhibit polyglutamine-induced androgen receptor aggregation. We identified D-enantiomeric peptide QF2D-2 (sqsqwstpqGkwshwprrr) as the most promising candidate. It binds to an androgen receptor fragment with 46 consecutive glutamine residues and decelerates its aggregation, even in seeded experiments. Therefore, QF2D-2 may be a promising drug candidate for SBMA treatment or even for all nine heritable polyglutamine diseases, since its aggregation-inhibiting property was shown also for a more general polyglutamine target.

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“…Besides the primary selection with 3CL pro as the target (Target Selection, TS), two control selections were performed as described before 40 diluted in washing buffer to a final concentration of 0.35 ng/µL, and 100 µL was added to each well for 1 h at RT. After 6 washing steps with 150 µL washing buffer, the supernatant was removed entirely, and 100 µL of the 3,3',5,5'-tetramethylbenzidine (TMB) solution (TMB was previously dissolved in 1 mL DMSO and diluted with 9 mL 0.05 M phosphate citrate buffer pH 5) were added into each well.…”

Section: Phage Displaymentioning

confidence: 99%

Discovery of all-D-peptide inhibitors of SARS CoV 2 3C-like protease

Eberle

Sevenich

Gering

et al. 2022

Preprint

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During the replication process of SARS-CoV-2 the main protease of the virus (3-chymotrypsin-like protease (3CLpro)) plays a pivotal role and is essential for the life cycle of the pathogen. Numerous studies have been conducted so far, which have confirmed 3CLpro as an attractive drug target to combat COVID-19. We describe a novel and efficient next generation sequencing (NGS) supported phage display selection strategy for the identification of a set of SARS-CoV-2 3CLpro targeting peptide ligands that inhibit the 3CL protease, in a competitive or non-competetive mode, in the low µM range. From the most efficient L-peptides obtained from the phage display, we designed all-D-peptides based on the retro-inverso (ri) principle. They had IC50 values also in the low µM range, and in combination even in the sub-micromolar range. The inhibition modes of these D-ri peptides were the same as their respective L-peptide versions. Our results demonstrate that retro-inverso obtained all-D-peptides interact with high-affinity and inhibit the SARS-CoV-2 3CL protease, thus reinforcing their potential as therapeutic agents. The here described D-ri peptides address limitations associated with current L-peptide inhibitors and are promising lead compounds. Further optimization regarding pharmacokinetic properties will allow the development of even more potent D-peptides to be used for the prevention and treatment of COVID-19.

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Ligand-Induced Stabilization of the Native Human Superoxide Dismutase 1

Cited by 10 publications

References 35 publications

Phage Display-Derived Compounds Displace hACE2 from Its Complex with SARS-CoV-2 Spike Protein

Phage Display-Derived Compounds Displace hACE2 from Its Complex with SARS-CoV-2 Spike Protein

Inhibition of Polyglutamine Misfolding with D-Enantiomeric Peptides Identified by Mirror Image Phage Display Selection

Discovery of all-D-peptide inhibitors of SARS CoV 2 3C-like protease

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