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A limulus intracellular coagulation inhibitor type 2. Purification, characterization, cDNA cloning, and tissue
Y Miura1, S Kawabata, Y Wakamiya
1Department of Biology, Faculty of Science, Kyushu University 33, Fukuoka, Japan.
The Journal of Biological Chemistry
|January 13, 1995
Summary
Horseshoe crab hemocytes contain a new serine protease inhibitor, LICI-2, which targets both horseshoe crab and mammalian proteases. This regulated secretory serpin is stored in granules and released upon stimulation.
Area of Science:
- Biochemistry
- Molecular Biology
- Immunology
Background:
- Previous identification of limulus intracellular coagulation inhibitor (LICI), a serine protease inhibitor in horseshoe crab hemocytes.
- LICI specifically inhibits limulus lipopolysaccharide-sensitive serine protease, factor C.
Purpose of the Study:
- To identify and characterize a novel serine protease inhibitor, designated LICI type-2 (LICI-2), from horseshoe crab hemocytes.
- To investigate the inhibitory activity, protein structure, gene expression, and localization of LICI-2.
Main Methods:
- Purification of LICI-2 from horseshoe crab hemocytes.
- Determination of inhibitory kinetics against various serine proteases.
- Isolation and sequencing of LICI-2 cDNA.
- Northern blotting for mRNA expression analysis.
- Immunoblotting to determine LICI-2 localization within hemocyte granules.
Main Results:
- LICI-2 inhibits horseshoe crab factor C and limulus clotting enzyme, as well as mammalian serine proteases like alpha-thrombin, kallikrein, plasmin, and tissue plasminogen activator.
- Purified LICI-2 is a 42 kDa glycoprotein; its cDNA codes for a 386 amino acid mature protein with a unique Lys-Ser reactive site.
- LICI-2 mRNA is exclusively expressed in hemocytes; the protein is stored in large granules and released upon stimulation.
Conclusions:
- LICI-2 represents a distinct intracellular serpin with broad inhibitory activity against both invertebrate and mammalian serine proteases.
- The unique reactive site and regulated secretory nature suggest a specialized role for LICI-2 in the horseshoe crab immune response.
- Proposed classification of LICIs into a new subfamily of regulated secretory intracellular serpins.
