Investigating the sequence properties that govern disorder and secretion of bacterial virulence proteins

Many proteins contain disordered segments that play important roles in vivo. Autotransporter (AT) proteins are key virulence proteins from pathogenic Gram-negative bacteria. AT proteins have disordered properties that have been shown to be crucial for their efficient secretion to the bacterial cell surface. We have extensively studied pertactin, a virulence protein secreted by Bordetella pertussis, the bacterium that causes whooping cough. Like several other ATs, the mature pertactin virulence protein contains two segments with distinctly different folding properties. In isolation, the more stable C-terminal segment (PCt) is stably folded whereas the N-terminal segment (PNt) is disordered. Our previous studies indicate that folding of PCt is important for the efficient secretion of pertactin. Moreover, we recently showed that local clustering of hydrophobic residues leads to a significant collapse of PNt in vitro. Interestingly, we found that the extent to which PNt adopts a disordered, highly expanded conformational ensemble regulates the efficient secretion of pertactin through the bacterial outer membrane. Taken together, these observations suggest that PNt contains sequence properties important for secretion. Because disordered properties are important for AT secretion, understanding the connections between amino acid sequence features, disordered conformations, and AT secretion will improve our understanding of virulence protein secretion. The present research project aims to build upon our recent findings to develop a predictive understanding of the sequence factors that regulate disorder in pertactin and other proteins. We are using a combination of biophysical and biochemical approaches to characterize the sequence determinants of disorder and test their impact on AT secretion. We expect that insight into the regulation of disorder in pertactin will likely also apply to other members of the AT protein family, the largest family of secreted virulence proteins in Gram-negative bacterial pathogens.