Design of a Cereblon construct for crystallographic and biophysical studies of protein degraders

Alena Kroupova, Valentina Spiteri, Zoe Rutter, Hirotake Furihata, Darren Darren, Sarath Ramachandran, Sohini Chakraborti, Kevin Haubrich, Julie Pethe, Denzel Gonzales, Andre Wijaya, Maria Rodriguez-Rios, Manon Sturbaut, Dylan Lynch, William Farnaby, Mark Nakasone, David Zollman (Lead / Corresponding author), Alessio Ciulli (Lead / Corresponding author)

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Abstract

The ubiquitin E3 ligase cereblon (CRBN) is the target of therapeutic drugs thalidomide and lenalidomide and is recruited by most targeted protein degraders (PROTACs and molecular glues) in clinical development. Biophysical and structural investigation of CRBN has been limited by current constructs that either require co-expression with the adaptor DDB1 or inadequately represent full-length protein, with high-resolution structures of degrader ternary complexes remaining rare. We present the design of CRBN midi, a construct that readily expresses from E. coli with high yields as soluble, stable protein without DDB1. We benchmark CRBN midi for wild-type functionality through a suite of biophysical techniques and solve high-resolution co-crystal structures of its binary and ternary complexes with degraders. We qualify CRBN midi as an enabling tool to accelerate structure-based discovery of the next generation of CRBN based therapeutics.

Original languageEnglish
Article number8885
Number of pages14
JournalNature Communications
Volume15
Issue number1
DOIs
Publication statusPublished - 15 Oct 2024

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