We examined the effects of a fragment of the substrate binding domain of DnaK on protein refolding from chemically denatured states. The fragment DnaK384-638, containing a full-length substrate binding domain, tightly binds to the unfolded protein in solution. The effects of DnaK384-638 on the reactivation of beta-galactosidase and luciferase were examined at low substrate concentration and low...

Chromatographic columns packed with commercially available hydrophobic interaction chromatography (HIC) media were found to be able to suppress aggregation and nevertheless had a tendency to promote the structural misfolding resulting in higher soluble protein recovery and lower specific activity than that by dilution when they were used to refold lysozyme, a model protein. Moreover, this misfo...

Recent advances in generating active proteins through refolding of bacterial inclusion body proteins are summarized in conjunction with a short overview on inclusion body isolation and solubilization procedures. In particular, the pros and cons of well-established robust refolding techniques such as direct dilution as well as less common ones such as diafiltration or chromatographic processes i...

Nekrasova et al. “Overexpression, Solubilization and puri fication of rat and human olfactory receptors' Eur. J. Bio chem. (1996) 238: 28-37.* Kiefer, H., et al. (1996) Expression of an Olfactory Receptor in Escherichia coli: Purification, Reconstitution, and Ligand Binding. Biochemistry 35:10677-16084. Kiefer, H., et al. (1999) Refolding of G protein-coupled receptors from inclusion bodies pro...

In metalloproteins, metal centers serve as active sites for a range of functional purposes and as important structural elements to facilitate protein folding and assembly. It is challenging to observe the reversible unfolding and refolding of metalloproteins because of a loss or decomposition of the metal center. Here, the reversible unfolding-refolding of the iron-sulfur protein rubredoxin was...

Protein disulphide isomerase (PDI) shows chaperone and anti-chaperone activities in assisting refolding of denatured and reduced lysozyme in redox Hepes buffer, but only chaperone activity in phosphate buffer and redox Hepes buffer containing 0.1 M NaCl. In non-redox Hepes buffer its anti-chaperone activity is very weak. PDI displays its anti-chaperone activity only for those substrates showing...

Trigger factor (TF) is a molecular chaperone that binds to bacterial ribosomes where it contacts emerging nascent chains, but TF is also abundant free in the cytosol where its activity is less well characterized. In vitro studies show that TF promotes protein refolding. We find here that ribosome-free TF stably associates with and rescues from misfolding a large repertoire of full-length protei...