Assembly of the Type II Secretion System such as Found in <em>Vibrio cholerae</em> Depends on the Novel Pilotin AspS

<div><p>The Type II Secretion System (T2SS) is a molecular machine that drives the secretion of fully-folded protein substrates across the bacterial outer membrane. A key element in the machinery is the secretin: an integral, multimeric outer membrane protein that forms the secretion pore. We show that three distinct forms of T2SSs can be distinguished based on the sequence characteristics of their secretin pores. Detailed comparative analysis of two of these, the <em>Klebsiella</em>-type and <em>Vibrio</em>-type, showed them to be further distinguished by the pilotin that mediates their transport and assembly into the outer membrane. We have determined the crystal structure of the novel pilotin AspS from <em>Vibrio cholerae</em>, demonstrating convergent evolution wherein AspS is functionally equivalent and yet structurally unrelated to the pilotins found in <em>Klebsiella</em> and other bacteria. AspS binds to a specific targeting sequence in the <em>Vibrio</em>-type secretins, enhances the kinetics of secretin assembly, and homologs of AspS are found in all species of <em>Vibrio</em> as well those few strains of <em>Escherichia</em> and <em>Shigella</em> that have acquired a <em>Vibrio</em>-type T2SS.</p>