The coiled-coil domains of the putative yeast cargo receptors Emp46p and Emp47p (Emp46p^cc and Emp47p^cc) assemble into heterocomplexes at neutral pH. Upon lowering the pH, the complex dissociates and reassembles into homo-oligomers. A glutamate residue (E303) located on the hydrophobic surface of Emp46p^cc serves as the pH-sensing switch for assembly and segregation, and we have suggested that its side chains are protonated in the heterocomplex, even at neutral pH. To examine this hypothesis, we constructed two structural models in which the side chains of E303 were negatively charged or protonated and analyzed the effects of these charged states on the structure of the heterocomplex using molecular dynamics (MD) simulations. The calculated structures suggested the side chains of E303 to be protonated in the heterocomplex, even at neutral pH. Based on these computational results, the pH dependence of Emp47p^cc homo-oligomer assembly was experimentally modified by a glutamate mutation on its hydrophobic surface. The Q306E mutant of Emp47p^cc underwent a structural transition at physiological pH. Our results suggest a method for modifying pH-dependent protein-protein interactions.

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Journal Article | Research Support, Non-U.S. Gov't

Authors

Kato K, Nakayoshi T, Kurimoto E, Oda A

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