SPI “sandwich”: Combined SUMO‐Peptide‐Intein expression system and isolation procedure for improved stability and yield of peptides

• DOI: 10.1002/pro.4316
• Publication Date: 2022-04-27T10:37:12Z
• Authors: Tess Lamer; Marco J. van Belkum; Anjalee Wijewardane; Sorina Chiorean; Leah A. Martin‐Vi...; John C. Vederas
• Keywords: Intein; Maltose-binding protein; Protein tag; Cecropin

Recombinant peptide production in Escherichia coli is often accomplished through cloning and expression of a fusion protein. The protein partner generally has two requirements: (a) it contains an affinity tag to assist with purification (b) can be cleaved off leave only the desired sequence behind. Common soluble partners include small ubiquitin-like modifier (SUMO), maltose-binding (MBP), glutathione S-transferase (GST), or intein proteins. However, heterologously expressed peptides suffer from proteolytic degradation instability. This pose major issue for applications requiring large amount purified peptide, such as NMR structural assignments biochemical assays. Improving yield by testing various isolation conditions requires significant effort may not lead improved results. Here, we cloned four different SUMO These (lactococcin A, leucocin faerocin MK, neopetrosiamide A) were truncated during fusions, resulting low yields peptide. To prevent this improve yield, designed new system create "sandwiched" form: His6 -SUMO-peptide-intein (SPI). sandwiched more stable protected against degradation, (up 17-fold) under set standard procedures. SPI uses commercially available vectors techniques, therefore offer economical facile route that undergo degradation.