The nerve growth factor. Amino acid composition and physicochemical properties.

Abstract

The nerve growth factor, purified from adult mouse submaxillary gland was analyzed for amino acid composition, isoelectric point and ultraviolet absorption spectrum. 1 The amino acid analysis showed the presence of ten-half cystine residues per mole of the nerve growth factor. Reagents for free sulfhydryl groups, such as 5,5-dithio-bis-(2-nitrobenzoic) acid, p-hydroxy-mercury benzoate and iodoacetamide did not react with the protein even in presence of 8 M urea or 6 M guanidine · HCl; this suggests that all of the half-cystine residues are present in the nerve growth factor molecule in the oxidized form. From the amino acid composition the weight average of nitrogen was estimated as 17.62% and the partial specific volume was calculated as 0.716 ml per g. 2 The ultraviolet absorption spectrum at pH 5.0 showed a maximum in the 280–282 nm region and a second peak at 289 nm; the latter may indicate that the tryptophanyl residues present in the molecule are rather exposed. The ultraviolet spectrum did not change in presence of 8 M urea. 3 The isoelectric point, determined by the electrofocusing technique, was 9.30. This value explains the poor solubility observed for the nerve growth factor at neutral and alkaline pH and it is in agreement with the experimental observation that pH 5.0 is more indicated for the recovery and maintainance of the nerve growth factor activity during the course of the purification.