Of linkers and autochaperones: an unambiguous nomenclature to identify common and uncommon themes for autotransporter secretion

Abstract

Autotransporter (AT) proteins provide a diverse array of important virulence functions to Gram-negative bacterial pathogens, and have also been adapted for protein surface display applications. The “autotransporter” moniker refers to early models that depicted these proteins facilitating their own translocation across the bacterial outer membrane. Although translocation is less autonomous than originally proposed, AT protein segments upstream of the C-terminal transmembrane β-barrel have nevertheless consistently been found to contribute to efficient translocation and/or folding of the N-terminal virulence region (the “passenger”). However, defining the precise secretion functions of these AT regions has been complicated by the use of multiple overlapping and ambiguous terms to define AT sequence, structural, and functional features, including “autochaperone”, “linker” and “junction”. Moreover, the precise definitions and boundaries of these features vary among ATs and even among research groups, leading to an overall murky picture of the contributions of specific features to translocation. Here we propose a unified, unambiguous nomenclature for AT structural, functional and conserved sequence features, based on explicit criteria. Applied to 16 well studied AT proteins, this nomenclature reveals new commonalities for translocation but also highlights that the autochaperone function is less closely coupled with a conserved sequence element than previously believed.

Multiple Driving Forces Required for Efficient Secretion of Autotransporter Virulence Proteins

“…This implies that longer passengers will be more difficult to secrete and may rely more on folding as a driving force. Consistent with this hypothesis, large endogenous AT passengers appear to have stricter requirements for efficient secretion than small heterologous passengers (6,51). For example, secretion of IcsA is compromised by mutations in the C-terminal end of the endogenous passenger (52), whereas substituting the entire passenger for the smaller heterologous MalE protein results in efficient secretion (43).…”

“…1) Equilibrium constants for folding were set to 10 6 . This is an order of magnitude typical for the stability of many proteins and corresponds to a ⌬G of ϳϪ8 kcal mol Ϫ1 .…”

“…1) (6). Each autotransporter includes an N-terminal signal sequence that directs transport across the inner membrane.…”

Looks can be deceiving: recent insights into the mechanism of protein secretion by the autotransporter pathway

“…All of the C‐terminal domains that have been crystallized to date form nearly superimposable 12‐stranded β‐barrels (Oomen et al ., ; Barnard et al ., ; van den Berg, ; Tajima et al ., ; Zhai et al ., ). The two major domains are connected by a short ‘linker’, part of which forms an α‐helical segment that is embedded inside the β‐barrel domain (Drobnak et al ., ,). In a recent survey, hidden Markov models identified > 1500 genes that likely encode autotransporters in the genomes of Proteobacteria, Chlamydiales and Fusobacteria and confirmed that the autotransporter pathway is very widespread (Celik et al ., ).…”

The passenger‐associated transport repeat promotes virulence factor secretion efficiency and delineates a distinct autotransporter subtype

“…Strikingly, ∼ 30% of the unique AT representatives analyzed contain a PATR. This is comparable with the abundance of the Pertactin‐like (PL) region, which we found here to be ∼ 37% (slightly more than previous estimates; Drobnak et al ., ). However, the actual usage of the PATR is much higher than 30% as it is commonly present in many copies per passenger.…”