Optimization of the Magnetic Recovery of Hits from One-Bead–One-Compound Library Screens

Mendes, Kimberly; Ndungu, John M.; Clark, Lorraine F.; Kodadek, Thomas

doi:10.1021/acscombsci.5b00090

Cited by 30 publications

(22 citation statements)

References 34 publications

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“…Recent efforts in the Kodadek lab have extensively characterized methods for on-bead screening that minimize the number of false positive hit beads, 35,36 a pervasive issue in screening OBOC libraries. By screening libraries with a theoretical redundancy of at least three-fold, they demonstrated that redundant hits were almost always true positive ligands.…”

Section: Resultsmentioning

confidence: 99%

Chromodomain Ligand Optimization via Target-Class Directed Combinatorial Repurposing

et al. 2016

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Efforts to develop strategies for small molecule chemical probe discovery against the readers of the methyl-lysine (Kme) post-translational modification have been met with limited success. Targeted disruption of these protein-protein interactions via peptidomimetic inhibitor optimization is a promising alternative to small molecule hit discovery; however, recognition of identical peptide motifs by multiple Kme reader proteins presents a unique challenge in the development of selective Kme reader chemical probes. These selectivity challenges are exemplified by the Polycomb repressive complex 1 (PRC1) chemical probe, UNC3866, which demonstrates sub-micromolar off-target affinity toward the non-PRC1 chromodomains CDYL2 and CDYL. Moreover, since peptidomimetics are challenging subjects for structure-activity relationship (SAR) studies, traditional optimization of UNC3866 would prove costly and time-consuming. Herein, we report a broadly applicable strategy for the affinity-based, target-class screening of chromodomains via the repurposing of UNC3866 in an efficient, combinatorial peptide library. A first-generation library yielded UNC4991, a UNC3866 analog that exhibits a distinct selectivity profile while maintaining sub-micromolar affinity toward the CDYL chromodomains. Additionally, in vitro pull-down experiments from HeLa nuclear lysates further demonstrate the selectivity and utility of this compound for future elucidation of CDYL protein function.

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Section: Resultsmentioning

confidence: 99%

Chromodomain Ligand Optimization via Target-Class Directed Combinatorial Repurposing

et al. 2016

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“…S60). These results highlight the importance of the magnetic bead selection and suggest that the protocol has yet to be optimized (70). Significantly higher enrichments can probably be achieved, since magnetic bead selection can enrich yeast cells 10 4 -fold with nearquantitative yield (69).…”

Section: A Two-stage Magnetic Bead Selection/on-bead Screen Improvesmentioning

confidence: 93%

Xenoprotein engineering via synthetic libraries

Gates

Vinogradov

Quartararo

et al. 2018

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

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Chemical methods have enabled the total synthesis of protein molecules of ever-increasing size and complexity. However, methods to engineer synthetic proteins comprising noncanonical amino acids have not kept pace, even though this capability would be a distinct advantage of the total synthesis approach to protein science. In this work, we report a platform for protein engineering based on the screening of synthetic one-bead one-compound protein libraries. Screening throughput approaching that of cell surface display was achieved by a combination of magnetic bead enrichment, flow cytometry analysis of on-bead screens, and high-throughput MS/MS-based sequencing of identified active compounds. Direct screening of a synthetic protein library by these methods resulted in the de novo discovery of mirror-image miniprotein-based binders to a ∼150-kDa protein target, a task that would be difficult or impossible by other means.

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“…Positive beads are picked up manually under a stereo-microscope or by automated sorting with a COPAS instrument [99]. If the target is a protein, it can be used to decorate magnetic nanoparticles, which in turn can be used as probes to detect and isolate positive beads with magnetic bead sorting [100,101]. After positive beads are isolated, chemical decoding can be performed directly with Edman microsequencing if coding tag consists of α-amino acids [102].…”

Section: Oboc Peptide Librarymentioning

confidence: 99%

Tumor-targeting peptides from combinatorial libraries

Liu

Xiao

et al. 2017

Advanced Drug Delivery Reviews

164

102

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Cancer is one of the major and leading causes of death worldwide. Two of the greatest challenges infighting cancer are early detection and effective treatments with no or minimum side effects. Widespread use of targeted therapies and molecular imaging in clinics requires high affinity, tumor-specific agents as effective targeting vehicles to deliver therapeutics and imaging probes to the primary or metastatic tumor sites. Combinatorial libraries such as phage-display and one-bead one-compound (OBOC) peptide libraries are powerful approaches in discovering tumor-targeting peptides. This review gives an overview of different combinatorial library technologies that have been used for the discovery of tumor-targeting peptides. Examples of tumor-targeting peptides identified from each combinatorial library method will be discussed. Published tumor-targeting peptide ligands and their applications will also be summarized by the combinatorial library methods and their corresponding binding receptors.

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Optimization of the Magnetic Recovery of Hits from One-Bead–One-Compound Library Screens

Cited by 30 publications

References 34 publications

Chromodomain Ligand Optimization via Target-Class Directed Combinatorial Repurposing

Chromodomain Ligand Optimization via Target-Class Directed Combinatorial Repurposing

Xenoprotein engineering via synthetic libraries

Tumor-targeting peptides from combinatorial libraries

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