Immunoblot analysis of sub-cellular localization of PrtV protein in <i>V</i>. <i>cholerae</i>.

<p>(A) Immunoblot analyses of cell fractions from <i>V</i>. <i>cholerae</i> wild type strain C6706 (lanes 1, 3, 5, and 7) and the <i>prtV</i> mutant (lanes 2, 4, 6, and 8) using anti-PrtV serum (upper panel), anti-Crp antiserum (middle panel), and anti-β-lactamase antiserum (lower panel). Lanes 1 and 2: whole cell lysates; lanes 3 and 4, cytoplasmic fractions; lanes 5 and 6, periplasmic fractions; lanes 7 and 8, culture supernatants. Asterisks indicate the 81 kDa PrtV protein (B) Proteinase K susceptibility assay. OMVs from <i>V</i>. <i>cholerae</i> wild type strain C6706 were treated with 0.5 μg ml<sup>-1</sup> of proteinase K (PK), 1% SDS and/or the proteinase K inhibitor PMSF (1 mM) as indicated. Samples were examined by immunoblot analysis using polyclonal anti-PrtV antiserum (upper panel). Lane 1: OMVs treated with only PK; lane 2: OMVs treated with SDS and PK; lane 3: OMVs treated with SDS, PMSF, and PK; lane 4: control OMVs. The same membrane was re-probed with OmpU antiserum as an internal control (lower panel).</p>