Secretion of human growth hormone in Bacillus subtilis using prepropeptide coding region of Bacillus amyloliquefaciens neutral protease gene

Abstract

Using a secretion vector plasmid pES150, which contained a region coding for promoter and prepropeptide of Bacillus amyloliquefaciens neutral protease gene (Honjo et al. (1985) J. Biotechnol. 2, 73–84), a hybrid plasmid pES150GH was constructed. The plasmid pES150GH possesses a mature human growth hormone gene following the prepropeptide coding region. Bacillus subtilis N325 harboring pES150GH produced growth hormone in the medium. The secreted hormone was indistinguishable immunologically from human pituitary growth hormone on double immunodiffusion test. The level of secretory production of growth hormone by B. subtilis reached 40–50 mg l −1 of the culture. The size of the hormone secreted by B. subtilis was approximately 23 K on SDS-PAGE. This value was rather large in comparison with that of natural growth hormone (22 K). It was observed that 99% of synthesized growth hormone was excreted into the medium and that the mode of secretion was co-translational. The secreted growth hormone (23 K) is suggested to be degraded to a 15 K form in the culture medium by proteolysis.