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. 2013 Dec 23;8(12):e84512. doi: 10.1371/journal.pone.0084512

Figure 3. Mutational Analysis of the BamA β-barrel.

Figure 3

Detection of BamA proteins carrying insertions within the β-barrel domain. N-terminal 6His tags were introduced into the Q441, Q466, D503, Y509, W546, Y574, L613, Q664, N666, A714, A770, Q789 and A799 insertion constructs cloned into pET17b (see Fig. 2). Total membranes were prepared from JWD3 cells, containing pET17b, pET17b/6hisbamA or pET17b carrying 6His insertion mutants, grown in the presence of arabinose. 1.6 µg of total membrane protein was subjected to Western blotting with anti-6His antiserum (top panel) and 4 µg of protein was analysed using SDS-PAGE and stained with Coomassie blue (bottom panel). The location of 6HisBamA proteins, OmpF, OmpC and OmpA are indicated. The location of each insertion with respect to the secondary structure detailed in Fig. 2 is also given.