Periplasmic Proteins Of E.coli Are Highly Resistant To Aggregation

Periplasmic proteins of Gram-negative bacteria like Escherichia coli are subjected to immediate affect of environmental fluctuation that may unfold proteins, due to the permeability of the outer membrane to small molecules. They are thus supposedly protected by certain molecular chaperones. Nevertheless, no homologues of typical molecular chaperones have so far been found in periplasm, and the recently reported chaperone activities of periplasmic protein disulfide isomerase (PDI) and peptidyl pro-lyl isomerase (PPI) seem to be too weak to satisfy such assumed needs. In an attempt to reveal whether periplasmic proteins exhibit certain unusual properties, we discovered that such proteins as a whole are highly resistant to aggregation under a wide variety of denaturing conditions like high temperature, low pH, organic solvent and heavy metal. Furthermore, in an effort to unveil the nature behind this phenomenon we purified and examined four prominent periplasmic proteins. It is shown that these proteins unfold at rather mild denaturing conditions monitored by far-UV CD and trasverse urea-gradient PAGE; they synchronous expose hydrophobic surfaces during such unfolding process detected by bis-ANS hydrophobic fluorescence probe; but they do hardly form complexes with a typical molecular chaperone at those unfolded, hydrophobic surfaces exposed state. Based on these observations, we propose that the periplasmic proteins have been evolved to resist the formation of aggregates when subjected to various denaturing conditions and molecular chaperones may thus not be needed in periplasm. We also strongly suggest that the recently reported chaperone acitivity of periplasmic PDI-ase and PPIase assayed by traditional substrate should be reexamined using periplasmic proteins as substrate due to the significant difference between those two classes of proteins, the possible chaperone fuctions of PDIase and PPIase in vivo should be reappraised.