Abstract: Human nerve growth factor (NGF) was cloned and engineered for expression in a baculovirus‐infected Spodoptera frugiperda (SF‐9) insect cell system. Culture supernatants contained 2–3 mg/L of recombinant human NGF. The human NGF produced by this system was purified to apparent homogeneity with a single‐step affinity chromatography procedure using a high‐affinity monoclonal antibody originally raised against murine NGF. The purification procedure yielded 1–2 mg of pure, human NGF per liter of culture supernatant; i.e., approximately 60% recovery of the human NGF originally released into the culture medium. Although the gene transacted into the SF‐9 cells coded for pro‐NGF, the NGF recovered after purification was > 95% fully processed, mature protein. The KD for the affinity of the pure, recombinant human NGF for NGF receptor in PC12 membranes is 0.20 ± 0.05 nM. Activation of neurite outgrowth in PC12 cells occurs with ED50 values of 85 ± 20 pM and 9.6 ± 1.5 pM for a 3‐day primary response and a 1‐day secondary response, respectively. The pure, recombinant human NGF also stimulates a significant increase in dopamine content of PC12 cells with an ED50 of 5.8 ± 2.7 pM. These binding and biological activation properties are consistent with values observed using murine NGF purified from sub‐maxillary glands.
|Original language||English (US)|
|Number of pages||7|
|Journal||Journal of Neurochemistry|
|State||Published - Mar 1991|
- Human nerve growth factor
ASJC Scopus subject areas
- Cellular and Molecular Neuroscience