The preparation and properties of nerve growth factor protein at alkaline pH

The nerve growth factor (NGF) subunit of 7S NGF was isolated by chromatography at high pH on QAE Sephadex. It has the same specific NGF activity as βNGF isolated at acid pH, showing that this activity is an intrinsic property of the subunit and is independent of the pathway of dissociation. Continued exposure of the NGF subunit to high pH resulted in an increase in the amount of the minor species β²NGF and the formation of a new species, β³NGF, of even lower isoelectric point. These two species and the original major species of the preparation, β¹, were isolated by isoelectric focusing. All three species had the same specific NGF activity, but differed in their ability to reform 7S NGF. The β² species was one fifth as competent as β¹, while β³ was unable to regenerate 7S NGF. Addition of α and γ subunits to β¹NGF decreased the amount of NGF protein required to produce one Biological Unit of activity in the bioassay, but had no effect when added to β³NGF. The interactions between the subunits in 7S NGF therefore determine, in part, the specific activity of the NGF subunit.