Driving Forces for Protein Secretion Across the Bacterial Outer Membrane

Abstract

Autotransporters are a large and diverse class of monomeric virulence proteins secreted from Gram-negative bacterial pathogens. Secretion across the outer membrane is facilitated by the C-terminal translocator domain that creates a pore in the outer membrane, through which the N-terminal passenger (the functional, extracellular part of the protein) exits the cell. However, there is no ATP in the periplasm nor an ion gradient across the outer membrane. It is therefore not clear where the free energy comes from to drive the transport of the passenger through the pore. Using a computational model of secretion kinetics, we show that the free energy of passenger folding could be used as a driving force for secretion, provided that the passenger does not fold prematurely in the periplasm. We have tested this model experimentally by reversibly stalling secretion and probing the periplasmic conformation of the stalled protein. Our results show that the passenger remains unfolded in the periplasm, confirming that the major requirement for coupling folding to secretion is met. In addition, preliminary data indicates that mutations that disrupt folding also prevent efficient secretion. Further work is in progress to experimentally characterize the interplay between autotransporter passenger folding and secretion.