A fine balance of hydrophobic-electrostatic communication pathways in a pH-switching protein. | Academic Article individual record
abstract

Allostery is the phenomenon of coupling between distal binding sites in a protein. Such coupling is at the crux of protein function and regulation in a myriad of scenarios, yet determining the molecular mechanisms of coupling networks in proteins remains a major challenge. Here, we report mechanisms governing pH-dependent myristoyl switching in monomeric hisactophilin, whereby the myristoyl moves between a sequestered state, i.e., buried within the core of the protein, to an accessible state, in which the myristoyl has increased accessibility for membrane binding. Measurements of the pH and temperature dependence of amide chemical shifts reveal protein local structural stability and conformational heterogeneity that accompany switching. An analysis of these measurements using a thermodynamic cycle framework shows that myristoyl-proton coupling at the single-residue level exists in a fine balance and extends throughout the protein. Strikingly, small changes in the stereochemistry or size of core and surface hydrophobic residues by point mutations readily break, restore, or tune myristoyl switch energetics. Synthesizing the experimental results with those of molecular dynamics simulations illuminates atomistic details of coupling throughout the protein, featuring a large network of hydrophobic interactions that work in concert with key electrostatic interactions. The simulations were critical for discerning which of the many ionizable residues in hisactophilin are important for switching and identifying the contributions of nonnative interactions in switching. The strategy of using temperature-dependent NMR presented here offers a powerful, widely applicable way to elucidate the molecular mechanisms of allostery in proteins at high resolution.

authors
publication outlet

Proc Natl Acad Sci U S A

author list (cited authors)
MacKenzie, D., Schaefer, A., Steckner, J., Leo, C. A., Naser, D., Artikis, E., ... Meiering, E. M.
publication date
2022
keywords
  • Hydrophobic And Hydrophilic Interactions
  • Allostery
  • Protozoan Proteins
  • Hydrogen-Ion Concentration
  • Microfilament Proteins
  • PH Dependence
  • Nmr Temperature Dependence
  • Signal Transduction
  • Genes, Switch
  • Myristoylation
  • Protein Binding
  • Protein Structure, Tertiary
  • Switch
  • Static Electricity
PubMed ID
35737838
identifier
638769SE
Digital Object Identifier (DOI)
start page
e2119686119
volume
119
issue
26