Contributions of the different extramembranous domains of the mechanosensitive ion channel MscL to its response to membrane tension

MscL is a mechanosensitive channel that is gated by tension in the membrane bilayer alone. It is a homooligomer of a protein comprising two transmembr ane segments connected by an external loop, with the NH2 and COOH termini l ocated in the cytoplasm, The contributions of the extramembranous domains o f the channel to its activity were investigated by specific proteolysis dur ing patch-clamp experiments. Limited proteolysis of the COOH terminus or th e NH2 terminus increased the mechanosensitivity of the channel without chan ging its conductance. Strikingly, after cleavage of the external loop of ea ch monomer, the channel was still functional, and its mechanosensitivity wa s increased dramatically, indicating that the loop acts as a spring that re sists the opening of the channel and promotes its closure when it is open. These results indicate that the integrity of most of the extramembranous do mains is not essential for mechanosensitivity, They suggest that these doma ins counteract the movement of the transmembrane helices to which they are connected, thus setting the level of sensitivity of the channel to tension.