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The kinetic and thermodynamic consequences of intrinsic disorder in protein-protein recognition are controversial. We address this by inducing one partner of the high-affinity colicin E3 rRNase domain-Im3 complex (K(d) ≈ 10(-12) M) to become an intrinsically disordered protein (IDP). Through a variety of biophysical measurements, we show that a single alanine mutation at Tyr507 within the hydrophobic core of the isolated colicin E3 rRNase domain causes the enzyme to become an IDP (E3 rRNase(IDP)). E3 rRNase(IDP) binds stoichiometrically to Im3 and forms a structure that is essentially identical to the wild-type complex. However, binding of E3 rRNase(IDP) to Im3 is 4 orders of magnitude weaker than that of the folded rRNase, with thermodynamic parameters reflecting the disorder-to-order transition on forming the complex. Critically, pre-steady-state kinetic analysis of the E3 rRNase(IDP)-Im3 complex demonstrates that the decrease in affinity is mostly accounted for by a drop in the electrostatically steered association rate. Our study shows that, notwithstanding the advantages intrinsic disorder brings to biological systems, this can come at severe kinetic and thermodynamic cost.

Original publication

DOI

10.1021/ja512607r

Type

Journal article

Journal

J Am Chem Soc

Publication Date

29/04/2015

Volume

137

Pages

5252 - 5255

Keywords

Colicins, Escherichia coli, Intrinsically Disordered Proteins, Models, Molecular, Point Mutation, Protein Binding, Protein Conformation, Protein Folding, Protein Interaction Mapping, Protein Interaction Maps, Protein Structure, Tertiary, Ribonucleases, Ribosomal Proteins, Thermodynamics