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.
MeSH terms
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Amino Acid Sequence
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Biotechnology / methods*
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Chromatography, Affinity
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Electrophoresis, Gel, Two-Dimensional
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Endodeoxyribonucleases / isolation & purification
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Endoribonucleases / isolation & purification
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Endotoxins / isolation & purification
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Exopeptidases / isolation & purification
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Histidine / metabolism
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Humans
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Interleukin-1beta / biosynthesis*
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Interleukin-1beta / chemistry
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Interleukin-1beta / isolation & purification
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Models, Molecular
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Molecular Sequence Data
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Oligopeptides / metabolism
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Recombinant Proteins / biosynthesis*
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Recombinant Proteins / chemistry
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Recombinant Proteins / isolation & purification
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Sequence Analysis, Protein
Substances
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Endotoxins
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His-His-His-His-His-His
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Interleukin-1beta
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Oligopeptides
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Recombinant Proteins
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Histidine
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Endodeoxyribonucleases
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Endoribonucleases
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Serratia marcescens nuclease
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Exopeptidases