Xenoprotein engineering via synthetic libraries

Gates, Zachary P.; Vinogradov, Alexander A.; Quartararo, Anthony J.; Bandyopadhyay, Anupam; Choo, Zi-Ning; Evans, Ethan D.; Halloran, Kathryn H.; Mijalis, Alexander J.; Mong, Surin K.; Simon, Mark D.; Standley, Eric A.; Styduhar, Evan D.; Tasker, Sarah Z.; Touti, Faycal; Weber, Jessica M.; Wilson, J. L.; Jamison, Timothy F.; Pentelute, Bradley L.

doi:10.1073/pnas.1722633115

Cited by 42 publications

(45 citation statements)

References 99 publications

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“…Numerous approaches to peptide design have been introduced to make AMPs less toxic for humans while maintaining or improving their potency to eliminate bacteria [11,42], e.g., rational design [119,120], combinatorial peptide libraries [75,121], high-throughput screening [122,123], database-guided approaches [124,125], structure-function-guided design [86,126,127], and molecular dynamics simulations [44]. Three major methods to improve AMP function have been described: (i) High-throughput screening can be used to identify potential AMPs [128][129][130].…”

Section: Conclusion and Future Perspectivesmentioning

confidence: 99%

Development and Challenges of Antimicrobial Peptides for Therapeutic Applications

2020

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More than 3000 antimicrobial peptides (AMPs) have been discovered, seven of which have been approved by the U.S. Food and Drug Administration (FDA). Now commercialized, these seven peptides have mostly been utilized for topical medications, though some have been injected into the body to treat severe bacterial infections. To understand the translational potential for AMPs, we analyzed FDA-approved drugs in the FDA drug database. We examined their physicochemical properties, secondary structures, and mechanisms of action, and compared them with the peptides in the AMP database. All FDA-approved AMPs were discovered in Gram-positive soil bacteria, and 98% of known AMPs also come from natural sources (skin secretions of frogs and toxins from different species). However, AMPs can have undesirable properties as drugs, including instability and toxicity. Thus, the design and construction of effective AMPs require an understanding of the mechanisms of known peptides and their effects on the human body. This review provides an overview to guide the development of AMPs that can potentially be used as antimicrobial drugs.

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Section: Conclusion and Future Perspectivesmentioning

confidence: 99%

Development and Challenges of Antimicrobial Peptides for Therapeutic Applications

2020

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“…DELs are typically limited to three or four varied positions, due to inefficiencies in the chemistry used for their assembly, and as such are more often categorized as libraries of small molecules. OBOC libraries can easily incorporate more varied positions, but OBOC screening is technically challenging and typically examines onlỹ 10 6 compounds 24 . Affinity selection-mass spectrometry (AS-MS) [25][26][27][28][29] represents an alternative strategy for target-based discovery of chemically accessed peptide binders.…”

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confidence: 99%

Ultra-large chemical libraries for the discovery of high-affinity peptide binders

et al. 2020

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High-diversity genetically-encoded combinatorial libraries (10 8 −10 13 members) are a rich source of peptide-based binding molecules, identified by affinity selection. Synthetic libraries can access broader chemical space, but typically examine only~10 6 compounds by screening. Here we show that in-solution affinity selection can be interfaced with nano-liquid chromatography-tandem mass spectrometry peptide sequencing to identify binders from fully randomized synthetic libraries of 10 8 members-a 100-fold gain in diversity over standard practice. To validate this approach, we show that binders to a monoclonal antibody are identified in proportion to library diversity, as diversity is increased from 10 6-10 8. These results are then applied to the discovery of p53-like binders to MDM2, and to a family of 3-19 nM-affinity, α/β-peptide-based binders to 14-3-3. An X-ray structure of one of these binders in complex with 14-3-3σ is determined, illustrating the role of β-amino acids in facilitating a key binding contact.

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“…Through the generation of this library of homogeneously modified proteins it was anticipated that the effect of amino acid sequence, and both the position and valency of sulfation, on the inhibition of thrombin and on anticoagulant activity could be determined. Significant interest is emerging in the generation of libraries of proteins for therapeutic discovery programs (28,29). We envisaged that our approach to generate a focused synthetic library of modified proteins would provide the means to uncover key structure-activity relationships akin to small molecule-based medicinal chemistry.…”

Section: Significancementioning

confidence: 99%

Rapid assembly and profiling of an anticoagulant sulfoprotein library

Watson

Ripoll‐Rozada

Lee

et al. 2019

Proc. Natl. Acad. Sci. U.S.A.

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Hematophagous organisms produce a suite of salivary proteins which interact with the host’s coagulation machinery to facilitate the acquisition and digestion of a bloodmeal. Many of these biomolecules inhibit the central blood-clotting serine proteinase thrombin that is also the target of several clinically approved anticoagulants. Here a bioinformatics approach is used to identify seven tick proteins with putative thrombin inhibitory activity that we predict to be posttranslationally sulfated at two conserved tyrosine residues. To corroborate the biological role of these molecules and investigate the effects of amino acid sequence and sulfation modifications on thrombin inhibition and anticoagulant activity, a library of 34 homogeneously sulfated protein variants were rapidly assembled using one-pot diselenide-selenoester ligation (DSL)-deselenization chemistry. Downstream functional characterization validated the thrombin-directed activity of all target molecules and revealed that posttranslational sulfation of specific tyrosine residues crucially modulates potency. Importantly, access to this homogeneously modified protein library not only enabled the determination of key structure–activity relationships and the identification of potent anticoagulant leads, but also revealed subtleties in the mechanism of thrombin inhibition, between and within the families, that would be impossible to predict from the amino acid sequence alone. The synthetic platform described here therefore serves as a highly valuable tool for the generation and thorough characterization of libraries of related peptide and/or protein molecules (with or without modifications) for the identification of lead candidates for medicinal chemistry programs.

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Xenoprotein engineering via synthetic libraries

Cited by 42 publications

References 99 publications

Development and Challenges of Antimicrobial Peptides for Therapeutic Applications

Development and Challenges of Antimicrobial Peptides for Therapeutic Applications

Ultra-large chemical libraries for the discovery of high-affinity peptide binders

Rapid assembly and profiling of an anticoagulant sulfoprotein library

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