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Molecular mechanism of hemolymph clotting system in limulus
Abstracts
bial peptides which have been isolated from the acid extract ofamoeboeytes of Tachypleustridentatus and L/mu/us po/yphemus. These isopeptides consist of 17-18 basic and hydrophobic amino acids with two S-S bonds. Tachyplesin isopeptides exhibited a wide variety of biological activities: 1) inhibition of LPS-mediated activation of Factor C in Limulus amoeboeyte lysate (LAL), 2) complex formation with LPS and acidic phospholipids, 3) neutralization of pyrogenicity of LPS, 4) hemolysis of LPS-sensitized erythrocytes, 5) broad antimicrobial spectrum against Gram positive and negative bacteria as well as some fungi, and 6) antiviral activity against vesicular stomatitis virus and human immunodeficiency virus. Chemically synthesized Taehyplesin isopeptides showed activities similar to the natural ones. 6. MOLECULAR MECHANISM OF HEMOLYMPH CLO'I'I'ING SYSTEM IN LIMULUS. T. Muta, T. Miyata, T. Tokunaga, T. Nakamura and S. lwanaga. Department of
Biology, Faculty of Science, Kyushu University, Fukuoka 812, Japan. The hemolyrnph clotting
III
7. THE AMINO ACID SEQUENCES OF MULTIPLE LECTINS OF BARNACLE MEGABALANUS ROSA. tC Muramoto and H. Kamiya. School of Fisheries Sciences, lO,tasato
University,SanrilaAIwate 022-01, Japan.
Although the presence of multiple leetins within one species favors the hypothesis that invertebrate leetins function as recognition factors in defense reactions, little effort has been directed toward the structural analysis of multiple lectins. We studied the amino acid sequences of multiple galactose-binding iectins isolated from the coelomic fluid of the acorn barnacle, Megabalanus rosa. They were composed of three different molecular species with respect to molecular weight, designated BRA-1 (Mr 330k), BRA-2 (Mr 140k) and BRA-3 (Mr 64k). BRA-3 was composed of four identical subunits (Mr 16k) of 138 amino acids with no carbohydrate chain. The subunits were cross-linked by disulfide bonds to form dimers at the carboxyl-terminal segment. Both BRA-1 and BRA-2were composed of identical subunits (Mr 22k) which were also crosslinked by interchain disulfide bonds, but at the amino-terminal segment. The subunit was a glycoprotein of 173 amino acids. BRA-2 and BRA-3 were distinct molecules in spite of a significant homology in their amino acid sequences. The amino acid sequences included some regions homologous to those in other invertebrate lectins, such as sea urchin and flesh fly lectins, and the calcium-dependent vertebrate lectins.
systems in Limulus are of great importance not only in hemostasis but also in the immune response. To elucidate the molecular mechanism of these systems, eDNA of zymogen factor C, an initiator for the coagulation cascade and highly sensitive to LPS, was cloned from Tachypleus tridentatus hemocyte eDNA library and then its base sequences were determined. Zymogen factor C consisted of a single chain of glycoprotein 8. EFFECTS OF INFECTION WITH ECHIwith a moleeularweight of 123,000. It contained NOSTOMA PARAENSEI ON MORPHOLOGY 25 residues of putative signal peptide and a total AND PHAGOCYTIC ACTIVITY OF HEof 994 residues with six potential glycosylation MOCYTES OF BIOMPHALARIA GLA. sites in its sequence. The most interesting finding BRATA. S. Noda and E. S. Loker*. Department was that factor C had five repeated units ("suhi of Medical Zoology, Faculty of Medicine, domains") with about 60 amino acid residues, Kagoshima University, Kagoshima 890, Japan, which showed a significant homology with a fam- and *Department of Biology, University of New ily of tandem repeats found in factor XIII b Mexico, Albuquerque, NM 87131, USA. Hesubunit, complement factor B, Clr, Cls, protein H mocytes from M-line Biomphalaria glabrata and C4bp. In addition, a Cys-rich, an EGF-like, a snails exposed to infection with Echinostoma hepatic lectin-like and pro-rich domains were paraensei for 1, 8 or 30 days were studied with also found in the NH2-terminal portion. The phase contrast optics. The relative proportions of COOH-terminal portion of factor C contained a five hemoeyte categories (fully-spread granulotypical serine protease domain, showing the cytes, partially-spread granulocytes, round cells, strongest homology with human a-thrombin. byalinocytes and other cell types) were relatively Based on this structural evidence, the Limulus unaffected by the parasite at one day post-expoclotting system appears to exhibit an evolutionary sure. At 8 days post-exposure, snails showed relationship between the mammalian comple- marked increases in the proportions of partiallyment and coagulation systems. spread granuloeytes and round cells. At 30 days post-exposure, hemoeyte composition was still
bial peptides which have been isolated from the acid extract ofamoeboeytes of Tachypleustridentatus and L/mu/us po/yphemus. These isopeptides consist of 17-18 basic and hydrophobic amino acids with two S-S bonds. Tachyplesin isopeptides exhibited a wide variety of biological activities: 1) inhibition of LPS-mediated activation of Factor C in Limulus amoeboeyte lysate (LAL), 2) complex formation with LPS and acidic phospholipids, 3) neutralization of pyrogenicity of LPS, 4) hemolysis of LPS-sensitized erythrocytes, 5) broad antimicrobial spectrum against Gram positive and negative bacteria as well as some fungi, and 6) antiviral activity against vesicular stomatitis virus and human immunodeficiency virus. Chemically synthesized Taehyplesin isopeptides showed activities similar to the natural ones. 6. MOLECULAR MECHANISM OF HEMOLYMPH CLO'I'I'ING SYSTEM IN LIMULUS. T. Muta, T. Miyata, T. Tokunaga, T. Nakamura and S. lwanaga. Department of
Biology, Faculty of Science, Kyushu University, Fukuoka 812, Japan. The hemolyrnph clotting
III
7. THE AMINO ACID SEQUENCES OF MULTIPLE LECTINS OF BARNACLE MEGABALANUS ROSA. tC Muramoto and H. Kamiya. School of Fisheries Sciences, lO,tasato
University,SanrilaAIwate 022-01, Japan.
Although the presence of multiple leetins within one species favors the hypothesis that invertebrate leetins function as recognition factors in defense reactions, little effort has been directed toward the structural analysis of multiple lectins. We studied the amino acid sequences of multiple galactose-binding iectins isolated from the coelomic fluid of the acorn barnacle, Megabalanus rosa. They were composed of three different molecular species with respect to molecular weight, designated BRA-1 (Mr 330k), BRA-2 (Mr 140k) and BRA-3 (Mr 64k). BRA-3 was composed of four identical subunits (Mr 16k) of 138 amino acids with no carbohydrate chain. The subunits were cross-linked by disulfide bonds to form dimers at the carboxyl-terminal segment. Both BRA-1 and BRA-2were composed of identical subunits (Mr 22k) which were also crosslinked by interchain disulfide bonds, but at the amino-terminal segment. The subunit was a glycoprotein of 173 amino acids. BRA-2 and BRA-3 were distinct molecules in spite of a significant homology in their amino acid sequences. The amino acid sequences included some regions homologous to those in other invertebrate lectins, such as sea urchin and flesh fly lectins, and the calcium-dependent vertebrate lectins.
systems in Limulus are of great importance not only in hemostasis but also in the immune response. To elucidate the molecular mechanism of these systems, eDNA of zymogen factor C, an initiator for the coagulation cascade and highly sensitive to LPS, was cloned from Tachypleus tridentatus hemocyte eDNA library and then its base sequences were determined. Zymogen factor C consisted of a single chain of glycoprotein 8. EFFECTS OF INFECTION WITH ECHIwith a moleeularweight of 123,000. It contained NOSTOMA PARAENSEI ON MORPHOLOGY 25 residues of putative signal peptide and a total AND PHAGOCYTIC ACTIVITY OF HEof 994 residues with six potential glycosylation MOCYTES OF BIOMPHALARIA GLA. sites in its sequence. The most interesting finding BRATA. S. Noda and E. S. Loker*. Department was that factor C had five repeated units ("suhi of Medical Zoology, Faculty of Medicine, domains") with about 60 amino acid residues, Kagoshima University, Kagoshima 890, Japan, which showed a significant homology with a fam- and *Department of Biology, University of New ily of tandem repeats found in factor XIII b Mexico, Albuquerque, NM 87131, USA. Hesubunit, complement factor B, Clr, Cls, protein H mocytes from M-line Biomphalaria glabrata and C4bp. In addition, a Cys-rich, an EGF-like, a snails exposed to infection with Echinostoma hepatic lectin-like and pro-rich domains were paraensei for 1, 8 or 30 days were studied with also found in the NH2-terminal portion. The phase contrast optics. The relative proportions of COOH-terminal portion of factor C contained a five hemoeyte categories (fully-spread granulotypical serine protease domain, showing the cytes, partially-spread granulocytes, round cells, strongest homology with human a-thrombin. byalinocytes and other cell types) were relatively Based on this structural evidence, the Limulus unaffected by the parasite at one day post-expoclotting system appears to exhibit an evolutionary sure. At 8 days post-exposure, snails showed relationship between the mammalian comple- marked increases in the proportions of partiallyment and coagulation systems. spread granuloeytes and round cells. At 30 days post-exposure, hemoeyte composition was still