ASSEMBLY MECHANISM OF THE OLIGOMERIC STREPTOLYSIN O PORE - THE EARLY MEMBRANE LESION IS LINED BY A FREE-EDGE OF THE LIPID-MEMBRANE AND IS EXTENDED GRADUALLY DURING OLIGOMERIZATION

Streptolysin O (SLO) is a bacterial exotoxin that binds to cell membra nes containing cholesterol and then oligomerizes to form large pores. Along with rings, are-shaped oligomers form on membranes, It has been suggested that each are represents an incompletely assembled oligomer and constitutes a functional pore, faced on the opposite side by a fre e edge of the lipid membrane. We sought functional evidence in support of this idea by using an oligomerization-deficient, nonlytic mutant o f SLO. This protein, which was created by chemical modification of a s ingle mutant cysteine (T250C) with N-(iodoacetaminoethyl)-1-naphthylan nine-5-sulfonic acid, formed hybrid oligomers with active SLO on membr anes. However, incorporation of the modified T250C mutant inhibited su bsequent oligomerization, so that the hybrid oligomers were reduced in size. These appeared as typical are lesions in the electron microscop e, They formed pores that permitted passage of NaCl and calcein but re stricted permeation of large dextran molecules. The data indicate that the SLO pore is formed gradually during oligomerization, implying tha t pores lined by protein on one side and an edge of free lipid on the other may be created in the plasma membrane. Intentional manipulation of the pore size may extend the utility of SLO as a tool in cell biolo gical experiments.