E. coli heat stable enterotoxin STa is an agonist of the membrane guanylate cyclase C whose endogenous ligands are the peptide hormones guanylin and uroguanylin. Whereas these peptides contain only two disulfide bonds, STa is stabilized by one additional disulfide bridge. We chemically synthesized the enterotoxin STh that originates from the E. coli strain found in humans, and we determined its...

Background: Pivotal roles of Nerve growth factor (NGF) in the development and survival of both neuronal and non-neuronal cells indicate its potential for the treatment of neurodegenerative diseases. However, investigation of NGF deficits in different diseases requires the availability of properly folded human b-NGF. In previous studies bacterial expression of hNGF demonstrated the feasibility o...

The malignant trophoblastic cell line JAR was used as a model system to study protein folding in intact cells. We have used this model previously to identify conformational intermediates in the production of an assembly-competent form of the human chorionic gonadotropin beta subunit (Ruddon, R. W., Krzesicki, R. F., Norton, S. E., Beebe, J. S., Peters, B. P., and Perini, F. (1987) J. Biol. Chem...

Disulfide bonds have only rarely been found in intracellular proteins. That pattern is consistent with the chemically reducing environment inside the cells of well-studied organisms. However, recent experiments and new calculations based on genomic data of archaea provide striking contradictions to this pattern. Our results indicate that the intracellular proteins of certain hyperthermophilic a...

The essential flavoenzyme Ero1p both creates de novo disulfide bonds and transfers these disulfides to the folding catalyst protein disulfide isomerase (PDI). The recently solved crystal structure of Ero1p, in combination with previous biochemical, genetic and structural data, provides insight into the mechanism by which Ero1p accomplishes these tasks. A comparison of Ero1p with the smaller fla...

We report the first example of a synthetic thiol-based compound that promotes oxidative protein folding upon 1-equivalent loading to disulfide bonds in client afford native...

8 3.1. Catalysis of oxidative protein folding 8 3.2 De novo formation of disulfide bonds in E. coli: the discovery of DsbA 9 3.3. DsbA is the most oxidizing disulfide catalyst 11 3.4. DsbB provides the periplasm with oxidizing power 15 3.5. Correcting wrong disulfide bonds in the periplasm: disulfide bond isomerization by DsbC 18 3.6. DsbD provides reducing equivalents in a highly oxidizing env...

The cysteines of the Escherichia coli periplasmic enzyme alkaline phosphatase, which are involved in disulfide bonds in the native enzyme, were found to be fully reduced when the protein was retained in the cytoplasm. Under these circumstances the cysteines remained reduced for at least several minutes after the synthesis of the protein was completed. This contrasted with the normally exported ...