Bacterial Protein Extraction Protocol

Protein extraction was carried out as described previously [19 (link),84 (link)]. In brief, an inoculation loop full of bacterial culture was harvested and the cells resuspended in 100 μL of a solution containing 50% acetonitrile (ACN; Merck, Darmstadt, Germany) and 1% aqueous trifluoroacetic acid (TFA; Acros Organics, NJ, US). After vortexing and centrifuging, the supernatant was treated with a solution of lysis buffer containing 60 mM Tris-HCl pH 7.5, 1% lauryl maltoside, 5 mM phenylmethanesulfonyl fluoride (PMSF) (Sigma, St. Louis, MO, US), and 1% dithiothreitol (DTT) (Sigma Chemical Co., US). The supernatant was then transferred to a fresh tube and the amount of protein was determined by the bicinchoninic acid method (Sigma Chemical Co., US). All experiments were performed in triplicates.