title: The pro-sequence facilitates folding of human nerve growth factor from Escherichia coli inclusion bodies.
abstract: Nerve growth factor (beta-NGF), a neurotrophin required for the development and survival of specific neuronal populations, is translated as a prepro-protein in vivo. While the presequence mediates translocation into the endoplasmic reticulum, the function of the pro-peptide is so far unknown. As the pro-sequences of several proteins are known to promote folding of the mature part, the renaturation behaviour of recombinant human beta-NGF pro-protein was compared to that of the mature form. Expression of rh-pro-NGF in Escherichia coli led to the formation of inclusion bodies (IBs). The presence of the covalently attached pro-sequence significantly increased the yield and rate of refolding with concomitant disulfide bond formation when compared to the in vitro refolding of mature NGF (rh-NGF). Physicochemical characterization revealed that rh-pro-NGF is a dimer. The pro-peptide could be removed by limited proteolysis with trypsin yielding biologically active, mature rh-NGF. Furthermore, rh-pro-NGF exhibited biological activity in the same concentration range as rh-NGF.
authors.affiliation: Institut für Biotechnologie, Martin-Luther-Universität Halle-Wittenberg, Halle/Saale, Germany.
authors.forename: A
authors.initials: A
authors.lastname: Rattenholl
authors.forename: H
authors.initials: H
authors.lastname: Lilie
authors.forename: A
authors.initials: A
authors.lastname: Grossmann
authors.forename: A
authors.initials: A
authors.lastname: Stern
authors.forename: E
authors.initials: E
authors.lastname: Schwarz
authors.forename: R
authors.initials: R
authors.lastname: Rudolph
mesh_terms: D004591:Electrophoresis, Polyacrylamide Gel; D018274:Electroporation; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D002479:Inclusion Bodies / Q000378:metabolism; D020932:Nerve Growth Factor / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D010957:Plasmids; D017510:Protein Folding; D011498:Protein Precursors / Q000737:chemistry*; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry; D014461:Ultracentrifugation
chemical_list: D011498:Protein Precursors; D011994:Recombinant Proteins; C061397:pro-nerve growth factor, human; D020932:Nerve Growth Factor