Purification and characterization of human interleukin-1β expressed in recombinant Escherichia coli

Authors


Correspondence to P. Wingfield, Biogen S.A., P.O. Box, CH-1211 Genève 24, Switzerland

Abstract

The high-level expression of human interleukin-1β in Escherichia coli is described. The protein contributes about 12% of the total cell protein and is associated with the soluble cytoplasmic fraction of the cell. A method for the purification of the protein is given which is based on anion- and cation-exchange chromatographies. The isolated protein, shown to be homogeneous by several analytical methods, has been characterized by amino acid analysis, N- and C-terminal sequence analysis and analytical centrifugation. The protein is biologically active as demonstrated by two different in vitro assays, namely, the mononuclear cell factor (IL-1/MCF) assay and lymphocyte activating factor (IL-1/LAF) assay. The specific activities determined with the IL-1/MCF and IL-1/LAF assays, are 2 × 107 and 4 × 107 units mg−1, respectively.

Abbreviations
IL-1β

interleukin-1β

SDS-PAGE

polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate

Nbs2

5,5′-dithiobis(2-nitrobenzoic acid)

IL-1/LAF assay

lymphocyte activating factor assay

IL-1/MCF

mononuclear cell factor assay

PGE2

prostaglandin E2

GLC-MS

combined gas-liquid chromatography/mass spectrometry

HPLC

high-pressure liquid chromatography

Ancillary

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