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Development of a robust HTRF assay with USP7 full length protein expressed in E. coli prokaryotic system for the identification of USP7 inhibitors by Yihui Song; Shu Wang; Min Zhao; Bin Yu is a Chemistry article available to read on EtoBox.

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Deubiquitinating enzyme ubiquitin-specific protease 7 (USP7) is a promising therapeutic target. Several USP7 inhibitors accommodated in the catalytic triad of USP7 have been reported with the aid of high-throughput screening (HTS) methods using USP7 catalytic domain truncation. However, the drawbacks of previously reported biochemical cleavage assays, including poor stability, fluorescence interference, time-consuming, expensive, more importantly the selectivity issue, have challenged the USP7-targeted drug discovery. In this work, we demonstrated the functional heterogeneity and essential role of different structural elements in the USP7 full activation, highlighting the necessity of USP7 full length in drug discovery. Apart from reported two pockets in the catalytic triad, five additional ligandable pockets were predicted based on the proposed USP7 full length models by AlphaFold and homology modelling. A reliable homogeneous time-resolved fluorescence (HTRF) HTS method was established based on the cleavage mechanism of USP7 towards the ubiquitin precursor UBA10. The USP7 full length protein was successfully expressed in the relatively cost-effective E. coli prokaryotic system an

Who reads Development of a robust HTRF assay with USP7 full length protein expressed in E. coli prokaryotic system for the identification of USP7 inhibitors?

Author

Yihui Song; Shu Wang; Min Zhao; Bin Yu

Publisher

Elsevier BV

Published

2023

Language

EN

Field

Chemistry (Physical Sciences)