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A major difference between "good" bacteria (such as the ones that reside in your gut and assist with food digestion) and "bad" bacteria (pathogenic strains that cause serious and/or fatal infectious diseases) is that "bad" bacteria express a wide variety of virulence proteins. Many virulence proteins are secreted (transported) out to the outer surface of the bacteria, where they assist the bacteria in the adhesion to, invasion of, and recruitment of nutrients from host cells. In fact, protein secretion is so important to virulence that Gram-negative bacterial pathogens (think contaminated spinach, Jack-in-the-Box hamburgers) have evolved no less than seven discrete mechanisms to secrete proteins across their double-walled envelope.

We study the mechanism of autotransporter (AT; also called Type Va) protein secretion in Gram-negative pathogens. AT proteins represent the largest family of outer membrane secreted virulence proteins in Gram-negative bacteria. The name "autotransporter" refers to the apparent ability of these proteins to transport themselves across the outer membrane in the absence of ATP, a proton gradient, or any obvious source of energy. AT proteins vary widely in size, sequence, and function, but we have shown that >97% contain a specific structural domain, a monomeric right-handed beta-helix. Moreover, we have shown that a portion of this beta-helix is more stable than the remainder of the protein, and this portion is the first part of the protein to cross the bacterial membrane. We have hypothesized that the folding properties of this portion of the beta-helix contribute to efficient outer membrane secretion. Intriguingly, AT beta-helices fold very slowly in vitro, which might mimic the environment encountered in the intra-membrane space (periplasm), where premature folding of the beta-helix could block transport across the outer membrane.

Besingi RN & Clark PL (2015) Extracellular protease digestion to evaluate membrane protein cell surface localization. Nature Protocols 10, 2074-2080. [PDF]

Drobnak I, Braselmann E & Clark PL (2015) Multiple driving forces required for efficient secretion of autotransporter virulence proteins. Journal of Biological Chemistry 290, 10104-10116. [PDF]

Drobnak I, Braselmann E, Chaney JL, Leyton D, Bernstein HD, Lithgow T, Luirink J, Nataro JP & Clark PL (2015) Of linkers and autochaperones: An unambiguous nomenclature to identify common and uncommon themes for autotransporter secretion. Molecular Microbiology 95, 1-16. [PDF]

Besingi RN, Chaney JL & Clark PL (2013) An alternative outer membrane secretion mechanism for an autotransporter protein lacking a C-terminal stable core. Molecular Microbiology 90, 1028-1045. [PDF]

Braselmann E & Clark PL (2012) Autotransporters: The cellular environment reshapes a folding mechanism to promote protein transport. Journal of Physical Chemistry Letters 3, 1063-1071. [PDF] [cover]

Renn JP, Junker M, Besingi RN, Braselmann E & Clark PL (2012) ATP-independent control of autotransporter virulence protein transport via the folding properties of the secreted protein.Chemistry & Biology 19, 287-296. [PDF] [cover]

Junker M & Clark PL (2010) Slow formation of aggregation-resistant beta-sheet folding intermediates. Proteins: Structure, Function & Bioinformatics 78, 812-824. [PDF]

Junker M, Besingi RN & Clark PL (2009) Vectorial transport and folding of an autotransporter virulence protein during outer membrane secretion. Molecular Microbiology 71, 1323-1332. [PDF]

Renn JP & Clark PL (2008) A conserved stable core structure in the passenger domain beta-helix of autotransporter virulence proteins. Biopolymers 89, 420-427. [PDF]

Junker M, Schuster C, McDonnell AV, Sorg KE, Finn MC, Berger B & Clark PL (2006) The pertactin beta-helix folding mechanism suggests common themes for secretion and folding of autotransporter proteins. Proceedings of the National Academy of Sciences USA 103, 4918-4923. [PDF]