MBP-ELIC expression levels were tested in four different growth media types: Luria Broth (LB), Terrific Broth (TB), and ZYM-5052 and ZYM-512 auto-induction media, as well as the 5 E. coli strains listed above. LB was purchased from Difco. TB was made in-house and consisted of 1.2% tryptone, 2.4% yeast extract, 0.4% glycerol, 17 mM KH2PO4, and 72 mM K2HPO4. The ZYM-5052 auto-induction medium was made as previously described [24 (link)]. ZYM-512 media was made using the same protocol, with the exception of an increase to 0.1% glucose from 0.05% glucose. The components used to make the media are as follows: tryptone, yeast extract, MgSO4, 0.2× trace metals, Na2HPO4, Na2SO4, KH2PO4, NH4Cl), glucose, lactose, and glycerol.
Extraction tests of MBP-ELIC fusion were performed with the following detergents, which were purchased from Anatrace unless otherwise noted: 1% SDS, 1% Sarcosine, 20 mM Dodecyl Maltoside-DDM, 40 mM Decyl Maltoside-DM, 1.5% Anzergent 3–10, 1.5% Anzergent 3–12, 1.5% Thesit (Sigma), 1.5% Triton X-100, 1.5% Anapoe C12E10, 1.5% Anapoe C10E9. From this pool, Sarcosine, DDM, Anzergent 3–12, and Triton X-100 were chosen due to the large quantities of protein extracted during the solubilization test. The hydrodynamic properties of the purified ELIC were assessed by Size Exclusion Chromatography (SEC) on a BioRad EnRich SEC 650 10×300 column.
Extraction tests of MBP-ELIC fusion were performed with the following detergents, which were purchased from Anatrace unless otherwise noted: 1% SDS, 1% Sarcosine, 20 mM Dodecyl Maltoside-DDM, 40 mM Decyl Maltoside-DM, 1.5% Anzergent 3–10, 1.5% Anzergent 3–12, 1.5% Thesit (Sigma), 1.5% Triton X-100, 1.5% Anapoe C12E10, 1.5% Anapoe C10E9. From this pool, Sarcosine, DDM, Anzergent 3–12, and Triton X-100 were chosen due to the large quantities of protein extracted during the solubilization test. The hydrodynamic properties of the purified ELIC were assessed by Size Exclusion Chromatography (SEC) on a BioRad EnRich SEC 650 10×300 column.