Bacillus subtilis MraY in detergent-free system of nanodiscs wrapped by styrene-maleic acid copolymers

PLoS One. 2018 Nov 5;13(11):e0206692. doi: 10.1371/journal.pone.0206692. eCollection 2018.


As an integral membrane protein, purification and characterization of phospho-N- acetylmuramyl- pentapeptide translocase MraY have proven difficult. Low yield and concerns of retaining stability and activity after detergent solubilization have hampered the structure-function analysis. The recently developed detergent-free styrene-maleic acid (SMA) co-polymer system offers an alternative approach that may overcome these disadvantages. In this study, we used the detergent free system to purify MraY from Bacillus subtilis. This allowed efficient extraction of MraY that was heterologously produced in Escherichia coli membranes into SMA-wrapped nanodiscs. The purified MraY embedded in these nanodiscs (SMA-MraY) was comparable to the micellar MraY extracted with a conventional detergent (DDM) with regard to the yield and the purity of the recombinant protein but required significantly less time. The predominantly alpha-helical secondary structure of the protein in SMA-wrapped nanodiscs was also more stable against heat denaturation compared to the micellar protein. Thus, this detergent-free system is amenable to extract MraY efficiently and effectively while maintaining the biophysical properties of the protein. However, the apparent activity of the SMA-MraY was reduced compared to that of the detergent-solubilized protein. The present data indicates that this is caused by a lower accessibility of the enzyme in SMA-wrapped nanodiscs towards its polyisoprenoid substrate.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacillus subtilis / enzymology*
  • Bacillus subtilis / genetics
  • Bacterial Proteins / chemistry
  • Bacterial Proteins / genetics
  • Bacterial Proteins / isolation & purification*
  • Biophysical Phenomena
  • Detergents
  • Enzyme Stability
  • Escherichia coli / genetics
  • Kinetics
  • Maleates
  • Micelles
  • Nanostructures
  • Polystyrenes
  • Protein Conformation, alpha-Helical
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / isolation & purification
  • Transferases / chemistry
  • Transferases / genetics
  • Transferases / isolation & purification*


  • Bacterial Proteins
  • Detergents
  • Maleates
  • Micelles
  • Polystyrenes
  • Recombinant Proteins
  • styrene-maleic acid polymer
  • Transferases
  • mraY protein, Bacteria

Grant support

EB received ZonMW ( grant (No. 205.100.008) from the Netherlands Organization for Scientific Research (NWO). JMD is supported by the seventh Framework Program of the European Union (Marie Curie Initial Training Network “ManiFold”, Grant No. 317371). SS is supported by the Foundation for Fundamental Research on Matter ( of NWO. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.