Production and comprehensive quality control of recombinant human Interleukin-1beta: a case study for a process development strategy.
ABSTRACT We describe an efficient strategy to produce high-quality proteins by using a single large IMAC chromatography column and enzymatic His-tag removal via the TAGZyme system in pilot scale. Numerous quality assays demonstrated a high purity of the final product, the human cytokine Interleukin-1beta (IL-1beta). The protein preparation was apparently free of host cell proteins, endotoxins, protease, and aggregates. The N-terminal amino acid sequence of IL-1beta was in full agreement with the natural mature form of IL-1beta. The homogeneity of the product was further shown by X-ray structure determination which confirmed the previously solved structure of the protein. We propose the applied workflow as a strategy for industrial production of protein-based biopharmaceuticals.
04/2012; , ISBN: 978-953-51-0544-2
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ABSTRACT: Histidine-rich peptides are commonly used in recombinant protein production as purification tags, allowing the one-step affinity separation of the His-tagged proteins from the extracellular media or cell extracts. Genetic engineering makes feasible the post-purification His-tag removal by inserting, between the tag and the main protein body, a target site for trans-acting proteases or a self-proteolytic peptide with regulatable activities. However, for technical ease, His tags are often not removed and the fusion proteins eventually used in this form. In this commentary, we revise the powerful biological properties of histidine-rich peptides as endosomolytic agents and as architectonic tags in nanoparticle formation, for which they are exploited in drug delivery and other nanomedical applications. These activities, generally unknown to biotechnologists, can unwillingly modulate the functionality and biotechnological performance of recombinant proteins in which they remain trivially attached.Microbial Cell Factories 12/2011; 10:101. · 3.55 Impact Factor