Reduced outer membrane permeability of Escherichia coli O157 : H7: Suggested role of modified outer membrane porins and theoretical function in resistance to antimicrobial agents

Outer membrane permeability of Escherichia coli O157:H7 was determined by a n in vivo kinetic model with the periplasmic enzyme alkaline phosphatase [M artinez et al. (1996) Biochemistry 35, 1179-1186]. p-Nitrophenyl phospate ( PNPP) substrate, added to intact bacteria, must diffuse through the outer m embrane to reach the enzyme. At low substrate concentration the bacterium w as in the perfectly reactive state where all molecules that entered the per iplasm were captured and converted to product. Transmembrane diffusion was rate limiting, and the permeability of the outer membrane was determined fr om kinetic properties. The O157:H7 strain grown at 30 degreesC showed one-s ixth the permeability of wildtype E. coli grown at 30 degreesC. Wild-type b acteria grown at greater than or equal to 37 degreesC show a physiological response with a shift in expression of outer membrane porins that lowered p ermeability to PNPP by approximately 70%. The O157:H7 strain did not displa y this temperature-sensitive shift in permeability even though a change in porin expression could be visualized by staining intensity of Omp F and Omp C on acrylamide gels. Altered behavior of the O157:H7 membrane was also in dicated by a several thousand-fold lower response to transformation relativ e to wild-type E. coli. Matrix-assisted laser desorption ionization time of Right mass spectrometry and electrospray ionization mass spectrometry conf irmed the expression of the Omp F and Omp C variants that are unique to E. coli O157:H7. This reduced outer membrane permeability can contribute to en hanced resistance of O157:H7 to antimicrobial agents.