Our solutions to this challenge were to reduce the temperature during expression far below what is normally used, include media components to tightly regulate expression levels, and prolong expression experiments over several days rather than a few hours or overnight. Although >95% of the expressed protein was still insoluble this gave ~2-3-fold increase in soluble expression. Our next steps were to identify conditions for Ni chelate and size exclusion chromatography, which were different to standard methods, to improve purity at these stages. When all of these steps were combined together, along with a large 20 litre E.coli culture, we were able to generate 20mg of >90% pure protein. The protein was confirmed to be covalently lipid modified by mass spectrometry analysis.