Expression and purification of ELP-intein-tagged target proteins in high cell density E. coli fermentation.
BACKGROUND
Elastin-like polypeptides (ELPs) are useful tools that can be used to non-chromatographically purify proteins. When paired with self-cleaving inteins, they can be used as economical self-cleaving purification tags. However, ELPs and ELP-tagged target proteins have been traditionally expressed using highly enriched media in shake flask cultures, which are generally not amenable to scale-up.


RESULTS
In this work, we describe the high cell-density expression of self-cleaving ELP-tagged targets in a supplemented minimal medium at a 2.5 liter fermentation scale, with increased yields and purity compared to traditional shake flask cultures. This demonstration of ELP expression in supplemented minimal media is juxtaposed to previous expression of ELP tags in extract-based rich media. We also describe several sets of fed-batch conditions and their impact on ELP expression and growth medium cost.


CONCLUSIONS
By using fed batch E. coli fermentation at high cell density, ELP-intein-tagged proteins can be expressed and purified at high yield with low cost. Further, the impact of media components and fermentation design can significantly impact the overall process cost, particularly at large scale. This work thus demonstrates an important advances in the scale up of self-cleaving ELP tag-mediated processes.
Department of Chemical Engineering, Princeton University, Princeton, New Jersey 08544, USA.
Baley A
BA
Fong
David W
DW
Wood
D002452:Cell Count; D004549:Elastin / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism*; D005285:Fermentation; D047668:Inteins / Q000235:genetics*; D010455:Peptides / Q000737:chemistry* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*
D010455:Peptides; D011993:Recombinant Fusion Proteins; D004549:Elastin