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A Colorimetric Assay that Specifically Measures Granzyme B Proteolytic Activity: Hydrolysis of Boc-Ala-Ala-Asp-S-Bzl
Published on: November 28, 2014
Granzyme B is a novel interleukin-18 converting enzyme
Youichi Omoto1, Keiichi Yamanaka, Kazuya Tokime
1Department of Dermatology, Mie University, Graduate School of Medicine, Tsu, Mie 514-8507, Japan.
Journal of Dermatological Science
|July 13, 2010
Summary
Granzyme B (GrB) converts inactive pro-interleukin-18 (proIL-18) into active IL-18, even in cells lacking caspase-1. This finding reveals GrB as a key enzyme in initiating IL-18 release and inflammation.
Area of Science:
- Immunology
- Cell Biology
- Enzymology
Background:
- Granzyme B (GrB) is known to induce apoptosis but its role in other biological events is unclear.
- Interleukin-18 (IL-18) is a pro-inflammatory cytokine produced as an inactive precursor (proIL-18).
- While caspase-1 processes proIL-18 in myeloid cells, its absence in non-hematopoietic cells suggests alternative processing mechanisms.
Purpose of the Study:
- To investigate if Granzyme B (GrB) can convert pro-interleukin-18 (proIL-18) into its biologically active form.
- To explore GrB's role in IL-18 processing within non-hematopoietic cells.
Main Methods:
- Recombinant proIL-18 was incubated with Granzyme B and analyzed via immunoblotting.
- Biological activity of GrB-cleaved proIL-18 was assessed using IFN-gamma assays.
- IL-18 processing and IFN-gamma induction were examined using extracts from Granzyme B-positive/caspase-1-negative human CD8+ T cells and normal human keratinocytes.
Main Results:
- Granzyme B cleaved proIL-18 into a fragment with identical sequence and biological activity to mature IL-18 processed by caspase-1.
- Extracts from human CD8+ T cells effectively cleaved proIL-18 into mature IL-18.
- IFN-gamma induction was observed in normal human keratinocytes treated with CD8+ T cell extracts.
Conclusions:
- Granzyme B is identified as a potent enzyme for converting pro-interleukin-18 (proIL-18) into mature IL-18.
- Secreted Granzyme B from cytotoxic T lymphocytes and/or NK cells may trigger IL-18 release from target cells.
- This mechanism suggests a role for Granzyme B in initiating inflammation through IL-18 activation.
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