Interleukin 1 beta (IL-1 beta) processing in murine macrophages requires a structurally conserved homologue of human IL-1 beta converting enzyme.
Open Access
- 1 March 1993
- journal article
- research article
- Published by Proceedings of the National Academy of Sciences in Proceedings of the National Academy of Sciences
- Vol. 90 (5), 1809-1813
- https://doi.org/10.1073/pnas.90.5.1809
Abstract
Murine interleukin 1 beta (IL-1 beta) convertase (mICE) was identified in cytosolic extracts of peritoneal exudate cells (PECs) and macrophage cell lines. mICE cleaves both the human and mouse IL-1 beta precursors (pIL-1 beta) at sites 1 and 2 but fails to cleave a human pIL-1 beta (Asp116 to Ala) mutant at site 2, indicating that Asp is required to the left of the scissile bond. Ac-Tyr-Val-Ala-Asp-amino-4-methyl coumarin, patterned after site 2 of human pIL-1 beta, is a fluorogenic substrate for mICE, while the tetrapeptide aldehyde Ac-Tyr-Val-Ala-Asp-CHO is a potent inhibitor (Ki = 3 nM) that prevents generation and release of mature IL-1 beta by PECs (IC50 = 7 microM). Cloning of a full-length 1.4-kb cDNA shows that mICE is encoded as a 402-aa proenzyme (p45) that can be divided into a prodomain (Met1-Asp122), followed by a p20 subunit (Gly123-Asp296), a connecting peptide (Ser297-Asp314), and a p10 subunit (Gly315-His402). At the amino acid level, p45, p20, and p10 are 62%, 60%, and 81% identical with human IL-1 beta convertase (hICE). The active site Cys284 lies within a completely conserved stretch of 18 residues; however, Ser289 in hICE, which aligns with the catalytic region of serine and viral cysteinyl proteases, is absent from mICE. Expression in Escherichia coli of a truncated cDNA encoding Asn119-His402 generated active enzyme, which was autocatalytically processed at three internal Asp-Xaa bonds to generate a p20 subunit (Asn119-Asp296) complexed with either p11 (Ala309-His402) or p10. Recombinant mICE cleaves murine pIL-1 beta accurately at the Asp117-Val118 bond. The striking similarities of the human and murine enzymes will make it possible to assess the therapeutic potential of hICE inhibitors in murine models of disease.Keywords
This publication has 17 references indexed in Scilit:
- Molecular Cloning of the Interleukin-1β Converting EnzymeScience, 1992
- Protease pro region required for folding is a potent inhibitor of the mature enzymeProteins-Structure Function and Bioinformatics, 1992
- A novel heterodimeric cysteine protease is required for interleukin-1βprocessing in monocytesNature, 1992
- The insulin A and B chains contain sufficient structural information to form the native moleculeTrends in Biochemical Sciences, 1991
- Identification of a monocyte specific pre-interleukin 1 beta convertase activity.Proceedings of the National Academy of Sciences, 1989
- Pro-sequence of subtilisin can guide the refolding of denatured subtilisin in an intermolecular processNature, 1989
- Activation of interleukin‐ 1β by a co‐induced proteaseFEBS Letters, 1989
- Expression of membrane interleukin 1 by fibroblasts transfected with murine pro-interleukin 1 alpha cDNA.Proceedings of the National Academy of Sciences, 1988
- The molecular evolution of genes and proteins: a tale of two serinesNature, 1988
- Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4Nature, 1970