One of the main problems in manufacturing polypeptides is that the isolation and sometimes production method varies from protein to protein, depending on their properties. We prepared the required BioBricks and introduced a procedure that streamlines and unifies manufacturing of polypeptides which results, in principle independently of their sequence, in high yield polypeptide production in bacteria. We demonstrated that peptide production of which in bacteria is difficult due to proteolytic degradation of small unstructured polypeptides, its toxicity to bacteria, such as in antimicrobial peptides and difficult isolation due to small size, can be successfully and in high yield produced as a fusion partner of insoluble ketosteroid isomerase (KSI). Further, acidic cleavage of an Asp-Pro dipeptide between KSI carrier and desired polypeptide is a simple and efficient way for releasing the preferred polypeptide. Since the KSI remains insoluble after cleavage, polypeptide can be easily isolated with HPLC.
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