Folding of barnase in the presence of the molecular chaperone SecB.

SecB is a molecular chaperone dedicated to interact exclusively with proteins destined for translocation across membranes. We find that SecB interacts with barnase during its folding in a similar manner to its interaction with GroEL. On mixing acid-denatured barnase with SecB in a stopped-flow spectrofluorimeter under conditions that favour refolding, we observe a series of fluorescence changes, corresponding to the binding of the denatured protein and the subsequent refolding of multiply and singly bound forms. The different phases were assigned using a combination of kinetics and mutant proteins. The refolding of barnase when bound to SecB is strongly retarded but never blocked. Multiply bound barnase is less tightly bound and refolds with a higher rate constant than singly bound barnase. Up to 4 mol of denatured barnase bind to 1 mol of tetrameric SecB.