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. 2014 Dec 8;207(5):615–626. doi: 10.1083/jcb.201404127

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© 2014 Hamilton et al.

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Figure 5. — Bypassing the holin: Tat-dependent rescue of chitinase secretion in a chiW mutant. (A) Experimental design. The JJH04w strain carries an in-frame deletion in chiW, encoding a holin-like membrane protein but retains an active Tat translocase in the inner membrane. A plasmid-encoded fusion between the S. marcescens SufI twin-arginine signal peptide and the S. marcescens ChiX protein will be targeted to the periplasm via Tat. N, N terminus; C, C terminus. (B) The JJH04w (ΔchiW) strain was transformed with empty vector (pBAD18), a vector encoding native ChiX (pBAD-ChiX), or a vector encoding the spSufI::ChiX fusion protein (pBAD-Tat-ChiX). All strains were grown aerobically in rich medium containing 0.02% (wt/vol) l-arabinose before being separated into culture supernatant (SN) and whole cell (WC) fractions. Samples were analyzed for the presence of ChiC and the periplasmic control protein (maltose binding protein [MBP]) by Western immunoblotting.