High conductance, stretch activated channels have been discovered in both Gram positive and Gram negative bacteria and located in the inner membrane. The conductances were of various sizes and different kinetics. It was not completely clear whether these varied conductances reflected the activity of different channels, or whether at least some of them arose from the cooperative behaviour of a limited number of pores. Inhibition of stretch induced activity by lanthanides was overcome by increasing the suction applied. The channels were inhibited again by increasing the concentration of lanthanides, suggesting variation in the mechanical properties of the membrane possessing an extremely high number of binding sites of varying affinity for the trivalents.
The inner membrane fraction of Escherichia coli, fused into giant liposomes and studied by patch clamp, revealed several other channels which were insensitive to the applied suction.
Work has been carried out on the regulation and modulation of channel activity by cytoplasmic components with a view to identifying their functional role. Further biochemical characterisation of the channels will be carried out using a traditional isolation approach involving solubilization and reconstitution of protein fractions (the most convenient functional assay being provided by the planar bilayer technique), leading to a study of the molecular biology of the channels.
Colicin and phage deoxyribonucleic acid (DNA) uptake (translocation) studies have allowed, several characteristics to be clearly defined, including the participation of Tol proteins. Work has also progressed in the characterization of the potassium transport system of E. coli and the investigation of the giant channels of mitochondria.
The newly discovered pores of the inner (cell) membrane of bacteria will be studied taking advantage of their ion-conducting properties. Their presence in several species, their conductance properties, kinetics, regulation and inhibition characteristics will be established, and their function defined. Isolation and chemical and genetic characterization will be attempted. The channel(s) involved in phage infection will be characterized. The possible channel formation in bacteria treated with aminoglycosides will be investigated.
Funding SchemeCSC - Cost-sharing contracts
10099 San Mauro Torinese Torino