Studies on Porins of Bacteria

Abstract

The effect of various plasmolysing agents on the resistance of Pseudomonas aeruginosa and Escherichia coli to rifampicin and bacitracin was investigated. It was found that the resistance of cells treated with 0.22 M NaC1 + 0.01 M MgCl2 was unaffected, whereas 0.3 M NaC1 or 0.5 M sucrose increased the sensitivity of the organisms to the antibiotics. It was determined that isomaltitol (OS 346) was identified by the maltose uptake system in E. coli after growth of the organism in nutrient broth. Isomaltotriitol (OS 508) and isomaltotetritol (OS 670) were not recognised. After growth of the organism in chemically defined media (CDM) in the absence of maltose, OS 346 was actively removed from the environment but at a reduced level. An in vivo double label assay for determining the size of the outer membrane pores in Gram-negative bacteria was designed. The assay was based on a comparison of the uptake into the periplasmic space of 3H labelled molecules from an isomaltitol series with that of 14C sucrose. The exclusion limit of E. coli pores was found to be approximately 700 d mw, confirming results obtained by other workers. The pores in P. aeruginosa were found to have an exclusion limit of approximately 360 d mw or less, in contrast to previous data. The effect of various growth environments on pore size and outer membrane protein composition was investigated. It was found that alterations in the growth environment of either organism did not significantly affect the outer membrane pore size. The nutritional richness of the growth media (CDM as compared with nutrient broth) affected the protein composition of the P. aeruginosa outer membrane, whereas the E. coli outer membrane proteins were unaffected by variation in the organism's CDM growth environment. The finding that P. aeruginosa has outer membrane pores with an exclusion limit less than that of E. coli is significant in that P. aeruginosa is unusually resistant to many antibiotics in comparison with other Gram-negative bacteria. This high resistance could be explained by the very low exclusion limit of the outer membrane pores.