Limulus intracellular coagulation inhibitor type 3. Purification, characterization, cDNA cloning, and tissue localization

Kishan Lal Agarwala, Shun Ichiro Kawabata, Yoshiki Miura, Yuka Kuroki, Sadaaki Iwanaga

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36 Citations (Scopus)

Abstract

We reported that limulus intracellular coagulation inhibitor type-1 (LICI-1) (Miura, Y., Kawabata, S., and Iwanaga, S. (1994) J. Biol. Chem. 269, 542-547) and LICI type-2 (LICI-2) (Miura, Y., Kawabata, S., Wakamiya, Y., Nakamura, T., and Iwanaga, S. (1995) J. Biol. Chem. 270, 558-565) found in the hemocyte lysate belong to the serpin family. The LICI-1 specifically inhibits limulus lipopolysaccharide-sensitive serine protease, factor C (k1 = 2.5 x 106 M-1 s-1), whereas LICI-2 inhibits preferentially limulus clotting enzyme (k1 = 4.3 x 105 M-1 s-1). In our ongoing studies on limulus serpin, we found another inhibitor, named LICI type-3 (LICI-3), which strongly inhibits (1,3)-β-D-glucan-sensitive serine protease, factor G (k1 = 3.9 x 105 M-1 s-1). Thus, the limulus hemolymph coagulation cascade is effectively regulated by at least the three endogenous serpins. LICI-3, newly identified in hemocytes, is a single chain glycoprotein with an apparent M(r) = 53,000, the largest one among known limulus serpins. A cDNA sequence for LICI-3 coded a mature protein of 392 amino acids, of which 68 residues were confirmed by peptide sequencing. LICI-3 showed significant sequence similarity to LICI-1 (45.8% identity) and LICI-2 (33.7% identity). LICI-3 contained a putative reactive site, -Arg-Ser-, distinct from that of LICI-2 (-Lys-Ser-) but the same as that of LICI-1. Expression of LICI-3 mRNA was detected only in hemocytes, and not in heart, brain, stomach, intestine, coxal gland, and skeletal muscle. Immunoblotting of the hemocyte-derived large and small granules with antiserum against LICI-3 suggested that it is stored specifically in large granules, as in the case of LICI-1 and LICI-2, and is released in response to external stimuli.

Original languageEnglish
Pages (from-to)23768-23774
Number of pages7
JournalJournal of Biological Chemistry
Volume271
Issue number39
DOIs
Publication statusPublished - 1996

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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