Secretion of mature Ifn–α2 and accumulation of uncleaved precursor by bacillus subtilis transformed with a hybrid α–Amylase signal sequence–ifn–α2 Gene

Bacillus subtilis was transformed with a pUB110–derived vector carrying the Bacillus amyloliquefaciens α–amylase promoter and signal sequence to which the coding sequence for mature human interferon–α2 (IFN) was accurately fused. The level of secreted IFN was at most 1–3% of that of α–amylase produced by B. subtilis transformed with the same vector containing the intact α–amylase gene. IFN was secreted at a 13–fold lower rate (on a weight basis) than α–amylase during the early stationary phase of growth, before protease activity became significant. mRNA levels for the two proteins, as determined by S₁ mapping, were similar. IFN–producing cells contained substantial amounts of a 21 kD protein, which was identified as a hybrid IFN precursor. The amount of insoluble mature and precursor IFN accumulating in the cells after 24 hours of incubation was 30–40 fold greater than the amount of IFN in the medium.