A hydrophilic microenvironment in the substrate-translocating groove of the YidC membrane insertase is essential for enzyme function

J Biol Chem. 2022 Mar;298(3):101690. doi: 10.1016/j.jbc.2022.101690. Epub 2022 Feb 9.

Abstract

The YidC family of proteins are membrane insertases that catalyze the translocation of the periplasmic domain of membrane proteins via a hydrophilic groove located within the inner leaflet of the membrane. All homologs have a strictly conserved, positively charged residue in the center of this groove. In Bacillus subtilis, the positively charged residue has been proposed to be essential for interacting with negatively charged residues of the substrate, supporting a hypothesis that YidC catalyzes insertion via an early-step electrostatic attraction mechanism. Here, we provide data suggesting that the positively charged residue is important not for its charge but for increasing the hydrophilicity of the groove. We found that the positively charged residue is dispensable for Escherichia coli YidC function when an adjacent residue at position 517 was hydrophilic or aromatic, but was essential when the adjacent residue was apolar. Additionally, solvent accessibility studies support the idea that the conserved positively charged residue functions to keep the top and middle of the groove sufficiently hydrated. Moreover, we demonstrate that both the E. coli and Streptococcus mutans YidC homologs are functional when the strictly conserved arginine is replaced with a negatively charged residue, provided proper stabilization from neighboring residues. These combined results show that the positively charged residue functions to maintain a hydrophilic microenvironment in the groove necessary for the insertase activity, rather than to form electrostatic interactions with the substrates.

Keywords: S. mutans; YidC; membrane biogenesis; membrane transport; molecular dynamics.

MeSH terms

  • Bacillus subtilis / enzymology
  • Cell Membrane / metabolism
  • Escherichia coli / chemistry
  • Escherichia coli / enzymology
  • Escherichia coli Proteins* / chemistry
  • Escherichia coli Proteins* / metabolism
  • Hydrophobic and Hydrophilic Interactions
  • Membrane Transport Proteins* / chemistry
  • Membrane Transport Proteins* / metabolism
  • Structure-Activity Relationship

Substances

  • Escherichia coli Proteins
  • Membrane Transport Proteins
  • YIDC protein, E coli