Abstract
The folding of human intestinal prolactase-phlorizin hydrolase (pro-LPH) has been analyzed in a cell-free transcription/translation system. In the presence of the thiol oxidant GSSG, disulfide bond formation in pro-LPH can be promoted concomitant with the binding of the molecule to a conformation-specific monoclonal anti-LPH antibody. Under these conditions, pro-LPH does not bind to the molecular chaperone BiP. In the absence of GSSG, on the other hand, pro-LPH does not bind to the monoclonal anti-LPH antibody, but can be immunoprecipitated with a polyclonal antibody that is directed against a denatured form of the enzyme. In this case, interaction of pro-LPH with immunoglobulin heavy chain binding protein can be discerned. The results demonstrate the existence of intramolecular disulfide bonds that are essential for the promotion of pro-LPH to a native conformation. Furthermore, BiP is involved in the folding events of pro-LPH.
Publication types
-
Research Support, Non-U.S. Gov't
MeSH terms
-
Adenosine Triphosphate / metabolism
-
Alkylation
-
Biopsy
-
Carrier Proteins / metabolism
-
Disulfides / metabolism*
-
Electrophoresis, Polyacrylamide Gel
-
Endoplasmic Reticulum Chaperone BiP
-
Enzyme Precursors / biosynthesis
-
Enzyme Precursors / chemistry*
-
Enzyme Precursors / genetics
-
Enzyme Precursors / metabolism
-
Glutathione / analogs & derivatives
-
Glutathione / pharmacology
-
Glutathione Disulfide
-
Heat-Shock Proteins*
-
Humans
-
Intestine, Small / enzymology
-
Intestines / enzymology*
-
Lactase-Phlorizin Hydrolase / biosynthesis
-
Lactase-Phlorizin Hydrolase / chemistry*
-
Lactase-Phlorizin Hydrolase / genetics
-
Lactase-Phlorizin Hydrolase / metabolism
-
Microvilli / enzymology
-
Molecular Chaperones / metabolism
-
Oxidation-Reduction
-
Protein Folding*
-
Recombinant Proteins / metabolism
-
Trypsin / pharmacology
Substances
-
Carrier Proteins
-
Disulfides
-
Endoplasmic Reticulum Chaperone BiP
-
Enzyme Precursors
-
Heat-Shock Proteins
-
Molecular Chaperones
-
Recombinant Proteins
-
Adenosine Triphosphate
-
Lactase-Phlorizin Hydrolase
-
Trypsin
-
Glutathione
-
Glutathione Disulfide