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bioRxiv · 10.1101/2021.11.27.470016

α-Helix stabilization by co-operative side chain charge-reinforced interactions to phosphoserine in a basic kinase-substrate motif

Batchelor, M.·Dawber, R. S.·Wilson, A. J.·Bayliss, R.
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Abstract

How cellular functions are regulated through protein phosphorylation events that promote or inhibit protein-protein interactions (PPIs) is key to understanding regulatory molecular mechanisms. Whilst phosphorylation can orthosterically or allosterically influence protein recognition, phospho-driven changes in the conformation of recognition motifs are less well explored. We recently discovered that clathrin heavy chain recognises phosphorylated TACC3 through a helical motif that, in the unphosphorylated protein, is disordered. However, it was unclear whether and how phosphorylation could stabilize a helix in a broader context. In the current manuscript, we address this challenge using poly-Ala based model peptides and a suite of circular dichroism and nuclear magnetic resonance spectroscopies. We show that phosphorylation of a Ser residue stabilizes the -helix in the context of an Arg(i - 3)pSeri Lys(i + 4) triad through charge-reinforced side chain interactions with positive co-operativity, whilst phosphorylation of Thr induces an opposing response. This is significant as it may represent a general method for control of PPIs by phosphorylation; basic kinase-substrate motifs are common with 55 human protein kinases recognising an Arg at a position -3 from the phosphorylated Ser, whilst the Arg(i - 3)pSeri Lys(i + 4) is a motif found in over 2000 human proteins.

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Batchelor, M., Dawber, R. S., Wilson, A. J., Bayliss, R.. 2021-11-27. α-Helix stabilization by co-operative side chain charge-reinforced interactions to phosphoserine in a basic kinase-substrate motif. https://doi.org/10.1101/2021.11.27.470016

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