3] High-level secretion of heterologous proteins Bacillus brevis

Abstract

Publisher Summary Among various host-vector systems for the production of foreign proteins in microorganisms, the use of Bacillus brevis ( B. brevis )as a host offers the advantage that proteins are secreted directly into the culture medium, where they are accumulated at high levels in a relatively pure state. B. brevis is isolated from soil as a protein-hyperproducing bacterium and is found to show little extracellular protease activity. Even when positive transformants of B. brevis are found, these cells sometimes cannot maintain the correct plasmid. Deletions of the foreign gene or cwp promoter are frequently found. Serial single-colony isolation of positive clones is helpful. The mutagenesis of B . brevis with N-methyl-N'-nitro-N-nitrosoguanidine (NTG) prior to transformation is also helpful to obtain clones that can maintain the plasmid with the correct structure. When clones maintaining the plasmid can be obtained but the amount of foreign proteins produced is not large, mutants producing the protein with improved yields can be isolated by the mutagenesis of the clones with NTG.