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.
Stenberg
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Cambridge Centre for Protein Engineering, Cambridge University Chemical Laboratory, Lensfield Road, Cambridge, CB2 1EW, UK.
Fersht
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D001407:Bacillus / Q000201:enzymology*; D001426:Bacterial Proteins / Q000378:metabolism*; D004926:Escherichia coli / Q000737:chemistry; D007700:Kinetics; D018832:Molecular Chaperones / Q000378:metabolism*; D009154:Mutation / Q000235:genetics; D011485:Protein Binding; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D012260:Ribonucleases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013050:Spectrometry, Fluorescence
D001426:Bacterial Proteins; D018832:Molecular Chaperones; C057842:SecB protein, Bacteria; D012260:Ribonucleases; C019760:Bacillus amyloliquefaciens ribonuclease