Albertsen, Louise4; Shaw, Allan C2; Norrild, Jens Chr.3; Strømgaard, Kristian4
1 Medicinal Chemistry Research, Department of Drug Design and Pharmacology, Faculty of Health and Medical Sciences, Københavns Universitet2 Novo Nordisk A/S3 Department of Organic Chemistry4 Medicinal Chemistry Research, Department of Drug Design and Pharmacology, Faculty of Health and Medical Sciences, Københavns Universitet
Peptides are of increasing interest as therapeutics in a wide range of diseases, including metabolic diseases such as diabetes and obesity. In the latter, peptide hormones such as peptide YY (PYY) and pancreatic peptide (PP) are important templates for drug design. Characteristic for these peptides is that they contain a C-terminal that is α-amidated, and this amidation is crucial for biological function. A challenge is to generate such peptides by recombinant means and particularly in a production scale. Here, we have examined an intein-mediated approach to generate a PYY derivative in a larger scale. Initially, we experienced challenges with hydrolysis of the intein fusion protein, which was reduced by a T3C mutation in the intein. Subsequently, we further engineered the intein to decrease the absolute size and improve the relative yield of the PYY derivative, which was achieved by substituting 54 residues of the 198 amino acid intein with an eight amino acid linker. The optimized intein construct was used to produce the PYY derivative under high cell density cultivation conditions, generating the peptide thioester precursor in good yields and subsequent amidation provided the target peptide.
Bioconjugate Chemistry, 2013, Vol 24, Issue 11, p. 1883-94