Proteome analsyis of Translocation phenomenon

Escherichia coli is the most widely used bacterial model organism and is the most commonly used host for the expression of recombinant proteins. Here we describe an unexpected protein translocation phenomenon in E. coli, whereby over-expression of recombinant proteins leads to high-level lysis-independent, mechanosensitive channel (MscL) dependent release of recombinant protein into the periplasmic space. Protein accumulation in the periplasm leads to protein release into the extracellular environment, independent of outer membrane vesicle formation. The translocated proteins retain their corresponding biological activity, and can be isolated directly from the extracellular medium with high purity and yield. Condition-specific metabolomic and proteomic analyses combined with statistical enrichment analysis indicate a role of both osmotic and translational stress responses in the regulation of the MscL-dependent translocation phenomenon. We suggest a model coupling translational stress to the regulation of MscL via the action of both osmotic stress and the Alternative ribosome-rescue factor A (ArfA) to explain this potentially very useful phenomenon.