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Neighboring phosphoSer‐Pro motifs in the undefined domain of IRAK 1 impart bivalent advantage for Pin1 binding
Author(s) -
Rogals Monique J.,
Greenwood Alexander I.,
Kwon Jeahoo,
Lu Kun Ping,
Nicholson Linda K.
Publication year - 2016
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.13943
Subject(s) - bivalent (engine) , chemistry , microbiology and biotechnology , computational biology , biophysics , biology , metal , organic chemistry
The peptidyl prolyl isomerase Pin1 has two domains that are considered to be its binding ( WW ) and catalytic ( PPI ase) domains, both of which interact with phosphorylated Ser/Thr‐Pro motifs. This shared specificity might influence substrate selection, as many known Pin1 substrates have multiple sequentially close phosphoSer/Thr‐Pro motifs, including the protein interleukin‐1 receptor‐associated kinase‐1 ( IRAK 1). The IRAK 1 undefined domain ( UD ) contains two sets of such neighboring motifs (Ser131/Ser144 and Ser163/Ser173), suggesting possible bivalent interactions with Pin1. Using a series of NMR titrations with 15N‐labeled full‐length Pin1 (Pin1‐ FL ), PPI ase, or WW domain and phosphopeptides representing the Ser131/Ser144 and Ser163/Ser173 regions of IRAK 1‐ UD , bivalent interactions were investigated. Binding studies using singly phosphorylated peptides showed that individual motifs displayed weak affinities (> 100 μ m ) for Pin1‐ FL and each isolated domain. Analysis of dually phosphorylated peptides binding to Pin1‐ FL showed that inclusion of bivalent states was necessary to fit the data. The resulting complex model and fitted parameters were applied to predict the impact of bivalent states at low micromolar concentrations, demonstrating significant affinity enhancement for both dually phosphorylated peptides (3.5 and 24 μ m for peptides based on the Ser131/Ser144 and Ser163/Ser173 regions, respectively). The complementary technique biolayer interferometry confirmed the predicted affinity enhancement for a representative set of singly and dually phosphorylated Ser131/Ser144 peptides at low micromolar concentrations, validating model predictions. These studies provide novel insights regarding the complexity of interactions between Pin1 and activated IRAK 1, and more broadly suggest that phosphorylation of neighboring Ser/Thr‐Pro motifs in proteins might provide competitive advantage at cellular concentrations for engaging with Pin1.