Open AccessThe pro‐sequence facilitates folding of human nerve growth factor from Escherichia coli inclusion bodies
Author(s) -
Rattenholl Anke,
Lilie Hauke,
Grossmann Adelbert,
Stern Anne,
Schwarz Elisabeth,
Rudolph Rainer
Publication year - 2001
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1046/j.1432-1327.2001.02232.x
Subject(s) - nerve growth factor , inclusion bodies , escherichia coli , endoplasmic reticulum , proteolysis , biochemistry , trypsin , chemistry , recombinant dna , protein folding , microbiology and biotechnology , peptide , biology , enzyme , gene , receptor
Nerve growth factor (β‐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 β‐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.