Mouse 7S nerve growth factor: complete sequence of a cDNA coding for the .alpha.-subunit precursor and its relationship to serine proteases

Abstract

Two synthetic oligonucleotides, one 14-mer and one 15-mer, each containing 32 sequences and corresponding to 2 regions of the partially determined protein sequence, were utilized to identify 3 cDNA [complementary DNA] clones coding for the precursor of the .alpha.-subunit of 7S mouse nerve growth factor (NGF). This library, containing 860 clones, had been preselected from a much larger one by low-stringency hybridization using a cDNA probe corresponding to one of the large family of glandular kallikreins expressed in the adult male mouse submandibular gland. Partial sequence analysis had previously established the .alpha.-subunit to be a member of this group, although with no demonstrable catalytic activity. Nucleotide sequence analysis of the longest of these clones (2A4) predicted the apparent complete amino acid sequence of the 265-residue precursor. One of the other clones (3F2) contained an A .fwdarw. G substitution at position 565 resulting in a Lys .fwdarw. Glu change at position 160 of the mature sequence. These clones probably represent 2 different alleles. Several amino acid changes, relative to other serine proteases, are evident which may account for the apparent lack of enzymatic activity. An Arg .fwdarw. Gln substitution at residue -1 would prevent cleavage of the putative activation peptide, and the deletion of residues 2-5 interrupts the highly conserved Ile/Val-Ile/Val-Gly-Gly N-terminal sequence. An Asp .fwdarw. Tyr substitution in the binding pocket and a Gly .fwdarw. His substitution near the active site serine also probably contribute to the inactive structure. The role of this subunit in NGF function remains obscure.