Biologically Active Ultra-Simple Proteins Reveal Principles of Transmembrane Domain Interactions

J Mol Biol. 2019 Sep 6;431(19):3753-3770. doi: 10.1016/j.jmb.2019.07.009. Epub 2019 Jul 10.

Abstract

Specific interactions between the helical membrane-spanning domains of transmembrane proteins play central roles in the proper folding and oligomerization of these proteins. However, the relationship between the hydrophobic amino acid sequences of transmembrane domains and their functional interactions is in most cases unknown. Here, we use ultra-simple artificial proteins to systematically study the sequence basis for transmembrane domain interactions. We show that most short homopolymeric polyleucine transmembrane proteins containing single amino acid substitutions can activate the platelet-derived growth factor β receptor or the erythropoietin receptor in cultured mouse cells, resulting in cell transformation or proliferation. These proteins displayed complex patterns of activity that were markedly affected by seemingly minor sequence differences in the ultra-simple protein itself or in the transmembrane domain of the target receptor, and the effects of these sequence differences are not additive. In addition, specific leucine residues along the length of these proteins are required for activity, and the positions of these required leucines differ based on the identity and position of the central substituted amino acid. Our results suggest that these ultra-simple proteins use a variety of molecular mechanisms to activate the same target and that diversification of transmembrane domain sequences over the course of evolution minimized off-target interactions.

Keywords: BPV E5 protein; BaF3 cells; erythropoietin receptor; helix; traptamer.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Amino Acid Sequence
  • Amino Acid Substitution
  • Animals
  • Cell Line
  • Humans
  • Mice
  • Mutagenesis / genetics
  • Mutation / genetics
  • Peptides / metabolism
  • Protein Domains
  • Protein Interaction Mapping*
  • Proteins / chemistry*
  • Proteins / metabolism*
  • Receptor, Platelet-Derived Growth Factor beta / chemistry
  • Receptor, Platelet-Derived Growth Factor beta / metabolism
  • Receptors, Erythropoietin / chemistry
  • Receptors, Erythropoietin / metabolism

Substances

  • Peptides
  • Proteins
  • Receptors, Erythropoietin
  • polyleucine
  • Receptor, Platelet-Derived Growth Factor beta