Target Identification Using Cell Permeable and Cleavable Chloroalkane Derivatized Small Molecules
In: Methods in Molecular Biology ISBN: 9781493972005; (2017)
Online
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Zugriff:
An important aspect for gaining functional insight into the activity of small molecules revealed through phenotypic screening is the identification of their interacting proteins. Yet, isolating and validating these interacting proteins remains difficult. Here, we present a new approach utilizing a chloroalkane (CA) moiety capture handle, which can be chemically attached to small molecules to isolate their respective protein targets. Derivatization of small molecules with the CA moiety has been shown to not significantly impact their cell permeability or potency, allowing for phenotypic validation of the derivatized small molecule prior to capture. The retention of cell permeability also allows for treatment of live cells with the derivatized small molecule and the CA moiety enables rapid covalent capture onto HaloTag coated magnetic beads. Additionally, several options are available for the elution of interacting proteins, including chemical cleavage of the CA moiety, competitive elution using excess unmodified small molecule, or sodium dodecyl sulfate (SDS) elution. These features taken together yield a highly robust and efficient process for target identification, including capture of weak or low abundance interactors.
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Target Identification Using Cell Permeable and Cleavable Chloroalkane Derivatized Small Molecules
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Autor/in / Beteiligte Person: | Urh, Marjeta ; Mendez‐Johnson, Jacqui ; Daniels, Danette L. ; Rachel Friedman Ohana |
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Quelle: | Methods in Molecular Biology ISBN: 9781493972005; (2017) |
Veröffentlichung: | Springer New York, 2017 |
Medientyp: | unknown |
ISBN: | 978-1-4939-7200-5 (print) |
DOI: | 10.1007/978-1-4939-7201-2_6 |
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