PT  - JOURNAL ARTICLE
AU  - Jamin B. Hein
AU  - Martha S. Cyert
AU  - Polly M. Fordyce
TI  - MRBLE-pep measurements reveal accurate binding affinities for B56, a PP2A regulatory subunit
AID  - 10.1101/2020.12.16.423088
DP  - 2020 Jan 01
TA  - bioRxiv
PG  - 2020.12.16.423088
4099  - http://biorxiv.org/content/early/2020/12/16/2020.12.16.423088.short
4100  - http://biorxiv.org/content/early/2020/12/16/2020.12.16.423088.full
AB  - Signal transduction pathways rely on dynamic interactions between protein globular domains and short linear motifs (SLiMs). The weak affinities of these interactions are essential to allow fast rewiring of signaling pathways and downstream responses, but pose technical challenges for interaction detection and measurement. We recently developed a technique (MRBLE-pep) that leverages spectrally encoded hydrogel beads to measure binding affinities between a single protein and 48 different peptide sequences in a single small volume. In prior work, we applied it to map the binding specificity landscape between calcineurin and the PxIxIT SLiM (Nguyen et al. 2019). Here, using peptide sequences known to bind the PP2A regulatory subunit B56, we systematically compare affinities measured by MRBLE-pep or isothermal calorimetry (ITC) and confirm that MRBLE-pep accurately quantifies relative affinity over a wide dynamic range while using a fraction of the material required for traditional methods such as ITC.Competing Interest StatementP.M.F. previously co-authored a patent describing production of spectrally encoded beads via ratiometric barcode lanthanide encoding (WO 2014/031902 A2).