Reconstitution of the Outer Membrane Protein OmpX in Nanodiscs

Abstract

Self-assembling bilayer phospholipid nanodiscs are a recently established tool when designing lipid-based nanodevices. The greater aim of nanodiscs is the stabilization of membrane proteins in the folded state, by incorporating them into nanodiscs. The outer membrane protein OmpX and the membrane scaffold protein MSP1D1-deltaH5, which is necessary for the assembling of the nanodics, are expressed in e. coli and later purified using established methods such as the affinity chromatography. Between different steps the solutions are run on SDS-polyacrylamide gels to check whether the protein is contained in the solution as expected. After both proteins have been purified and concentrated the nanodiscs are assembled with the self-assembly of the phospholipid bilayer nanodiscs. After the nanodics have been assembled, further experiments like size-exclusion-chromatography, circular dichroism spectroscopy and NMR spectroscopy are being done in order to get more data about the nanodisds and the protein OmpX. The purpose of assembling nanodiscs is to be able to carry out further structural determination more precisely.