Biochemical Properties and cDNa Cloning of Two New Lectins from the Plasma of Tachypleus tridentatus

Abstract

A Sepharose CL-4B-binding protein,Tachypleus plasma lectin 1 (TPL-1), and a lipopolysaccharide (LPS)-binding protein, Tachypleus plasma lectin-2 (TPL-2), have been isolated from the plasma ofTachypleus tridentatus and biochemically characterized. Each protein is coded by a homologous family of multigenes. TPL-1 binds to Sepharose CL-4B and was eluted with buffer containing 0.4m GlcNAc. The deduced amino acid sequence of TPL-1 consisted of 232 amino acids with an N-glycosylation site, Asn-Gly-Ser at residues 74–76. It shares a 65% sequence identity and similar internal repeats of about 20 amino acid motifs with tachylectin-1. Tachylectin-1 was identified as a lipopolysaccharide-agarose binding nonglycosylated protein from the amebocytes of T. tridentatus. TPL-2 was eluted from the LPS-Sepharose CL-4B affinity column in buffer containing 0.4m GlcNAc and 2 m KCl. The deduced amino acid sequence of TPL-2 consisted of 128 amino acids with anN-glycosylation site, Asn-Cys-Thr, at positions 3–5. It shares an 80% sequence identity with tachylectin-3, isolated from the amebocytes of T. tridentatus. TPL-2 purified by LPS-affinity column from the plasma predominantly exists as a dimer of a glycoprotein with an apparent molecular mass of 36 kDa. Tachylectin-3 is an intracellular nonglycosylated protein that also exists as a dimer in solution with an apparent molecular mass of 29 kDa. It recognizes Gram-negative bacteria through the 0-antigen of LPS. Western blot analyses showed that, in the plasma, TPL-1 and TPL-2 exist predominantly as oligomers with molecular masses above 60 kDa. They both bind to Gram-positive and Gram-negative bacteria, and this binding is inhibited by GlcNAc. Possible binding site of TPL-1 and TPL-2 to the bacteria could be at the NAc moiety of GlcNAc-MurNAc of the peptidoglycan. The physiological function of TPL-1 and TPL-2 is most likely related to their ability to form a cluster of interlocking molecules to immobilize and entrap invading organisms. A Sepharose CL-4B-binding protein,Tachypleus plasma lectin 1 (TPL-1), and a lipopolysaccharide (LPS)-binding protein, Tachypleus plasma lectin-2 (TPL-2), have been isolated from the plasma ofTachypleus tridentatus and biochemically characterized. Each protein is coded by a homologous family of multigenes. TPL-1 binds to Sepharose CL-4B and was eluted with buffer containing 0.4m GlcNAc. The deduced amino acid sequence of TPL-1 consisted of 232 amino acids with an N-glycosylation site, Asn-Gly-Ser at residues 74–76. It shares a 65% sequence identity and similar internal repeats of about 20 amino acid motifs with tachylectin-1. Tachylectin-1 was identified as a lipopolysaccharide-agarose binding nonglycosylated protein from the amebocytes of T. tridentatus. TPL-2 was eluted from the LPS-Sepharose CL-4B affinity column in buffer containing 0.4m GlcNAc and 2 m KCl. The deduced amino acid sequence of TPL-2 consisted of 128 amino acids with anN-glycosylation site, Asn-Cys-Thr, at positions 3–5. It shares an 80% sequence identity with tachylectin-3, isolated from the amebocytes of T. tridentatus. TPL-2 purified by LPS-affinity column from the plasma predominantly exists as a dimer of a glycoprotein with an apparent molecular mass of 36 kDa. Tachylectin-3 is an intracellular nonglycosylated protein that also exists as a dimer in solution with an apparent molecular mass of 29 kDa. It recognizes Gram-negative bacteria through the 0-antigen of LPS. Western blot analyses showed that, in the plasma, TPL-1 and TPL-2 exist predominantly as oligomers with molecular masses above 60 kDa. They both bind to Gram-positive and Gram-negative bacteria, and this binding is inhibited by GlcNAc. Possible binding site of TPL-1 and TPL-2 to the bacteria could be at the NAc moiety of GlcNAc-MurNAc of the peptidoglycan. The physiological function of TPL-1 and TPL-2 is most likely related to their ability to form a cluster of interlocking molecules to immobilize and entrap invading organisms. lipopolysaccharide C-reactive protein high performance liquid chromatography phosphocholine phosphoethanolamine polymerase chain reaction rapid amplification of cDNA ends polyacrylamide gel electrophoresis gene-specific primer tachylectin Tachypleus plasma lectin phenylthiohydantoin bovine serum albumin The innate and the adaptive immunities are the two general systems that mediate resistance to infectious agents. Although a certain form of adaptive immunity is present in all vertebrates, the invertebrates have developed only the innate immune system that has been thought of as an evolutionary rudiment, whose only function is to limit infection until adaptive immune response is induced. Recent studies have shown that the innate immune system has the capacity to induce costimulatory signals necessary for the activation and differentiation of lymphocytes (1Fearon D.T. Locksley R.M. Science. 1996; 272: 50-54Crossref PubMed Scopus (1431) Google Scholar, 2Lemaitre B. Nicolas E. Michaut L. Reichhart J.M. Hoffmann J.A. Cell. 1996; 86: 973-983Abstract Full Text Full Text PDF PubMed Scopus (2983) Google Scholar, 3Carroll M.C. Prodeus A.P. Curr. Opin. Immunol. 1998; 10: 36-40Crossref PubMed Scopus (188) Google Scholar). This finding has renewed interest on the studies of invertebrate and vertebrate innate immunology. The innate immune system uses germline-encoded receptors for recognition of common antigens on the surface of microbial pathogens. This feature distinguishes the innate immune system found in invertebrates from the adaptive immune system of the vertebrates that possess a repertoire of specific antigen receptors and antibodies. The conserved constituents or patterns, displayed by microorganisms, are recognized by pattern recognition molecules or receptors (4Medzhitov R. Janeway Jr., C.A. Cell. 1997; 91: 295-298Abstract Full Text Full Text PDF PubMed Scopus (1965) Google Scholar). These patterns, called pathogen-associated molecular patterns, seem to be shared among groups of pathogens. The lipopolysaccharides (LPS)1 of Gram-negative bacteria, lipoteichoic acid of Gram-positive bacteria, glycolipids of mycobacterium, and mannans of yeast are some examples. The innate defense system is designed to recognize those pathogen-associated molecular patterns. The horseshoe crab, an arthropod, has evolved only a nonclonal, or innate, defense system. The hemolymph and the hemocytes carry this defense system. Whereas the hemocytes, also named amebocytes, contain large and small granules that are filled with defense molecules, such as coagulation factors (5Muta T. Iwanaga S. Curr. Opin. Immunol. 1996; 8: 41-47Crossref PubMed Scopus (228) Google Scholar, 6Iwanaga S. Kawabata S. Muta T. J. Biochem. ( Tokyo ). 1998; 123: 1-15Crossref PubMed Scopus (253) Google Scholar, 7Fortes-Dias C.L. Minetti C.A.S. Lin Y. Liu T.-Y. Comp. Biochem. Physiol. 1993; 105: 79-85Crossref Scopus (24) Google Scholar), protease inhibitors (8Minetti C.S.A. Lin Y. Cislo T. Liu T.-Y. J. Biol. Chem. 1991; 266: 20773-20780Abstract Full Text PDF PubMed Google Scholar), and antimicrobial peptides (6Iwanaga S. Kawabata S. Muta T. J. Biochem. ( Tokyo ). 1998; 123: 1-15Crossref PubMed Scopus (253) Google Scholar), the hemoplymph contains three major proteins: hemocyanin, C-reactive proteins (CRPs), and α2-macroglobulin. Hemocyanin functions as an oxygen-carrying protein. CRPs are lectins that bind to phosphocholine of the pneumococcus C-polysaccharide (9Volanakis J.E. Kaplan M.H. Proc. Exp. Biol. Med. 1971; 136: 612-614Crossref PubMed Scopus (323) Google Scholar) and to the chromatin of damaged cells (10Robey F.A. Liu T.-Y. J. Biol. Chem. 1981; 256: 969-975Abstract Full Text PDF PubMed Google Scholar). α2-Macroglobulin exhibits protease inhibitory activity with a broad specificity that can block the activities of proteases secreted from invading microorganisms (11Enghild J.J. Thogersen I.B. Salvesen G. Fey G.H. Figler N.L. Gonias S.L. Pizzo S.V. Biochemistry. 1990; 29: 10070-10080Crossref PubMed Scopus (79) Google Scholar). TheLimulus CRPs, along with the C3 homologue, α2-macroglobulin, participate in a complement-like hemolytic activity in horseshoe crab hemolymph. Several lectins with a broad range of specificity have been identified in the amebocytes of horseshoe crab (12Liu T. Lin Y. Cislo T. Minetti C.A.S.A. Baba J.M.K. Liu T.Y. J. Biol. Chem. 1991; 266: 14813-14821Abstract Full Text PDF PubMed Google Scholar, 13Kawabata S. Iwanaga S. Dev. Comp. Immunol. 1999; 23: 391-400Crossref PubMed Scopus (87) Google Scholar). These lectins have been proposed to function in concert to defend horseshoe crabs from invading pathogens. However, since these lectins are present mostly in the granules of the hemocytes, they are unlikely to be involved in the immediate-early response of host-pathogen interaction. The plasma of horseshoe crab also contains lectin-like innate defense molecules (14Gokudan S. Muta T. Tsuda R. Koori K. Kawahara T. Seki N. Mizunoe Y. Wai S.N. Iwanaga S. Kawabata S. Proc. Natl. Acad. Sci. U. S. A. 1999; 96: 10086-10091Crossref PubMed Scopus (189) Google Scholar, 15Chiou S.T. Chen Y.W. Chen S.C. Chao C.F. Liu T.Y. J. Biol. Chem. 2000; 275: 1630-1634Abstract Full Text Full Text PDF PubMed Scopus (30) Google Scholar). In the previous study from this laboratory, we described the isolation and characterization of proteins that bind to Sepharose CL-4B, lipopolysaccharide of Escherichia coli, and protein A of Staphylococcus aureus from the plasma ofTachypleus tridentatus (15Chiou S.T. Chen Y.W. Chen S.C. Chao C.F. Liu T.Y. J. Biol. Chem. 2000; 275: 1630-1634Abstract Full Text Full Text PDF PubMed Scopus (30) Google Scholar). In the present study, we report biochemical characterization and cDNA cloning of two of the proteins, the Sepharose CL-4B-binding protein (TPL-1) and the lipopolysaccharide-binding protein (TPL-2), which we believe are involved in the innate immunity of horseshoe crabs. E. coli O55:B5 LPS was purchased from Sigma. Sepharose CL-4B, CNBr-activated Sepharose CL-4B, molecular weight standards, and staphylococcal protein A-Sepharose CL-4B were from Amersham Pharmacia Biotech (Uppsala, Sweden). Trypsin and complete protease inhibitor tablets were from Roche Molecular Biochemicals. Streptavidin-agarose and EZ-Link NHC-LC-Biotin were from Pierce. All other chemicals were of the highest quality commercially available. T. tridentatus were on the of crabs were by and hemolymph was in a containing of with 2 and protease inhibitor tablets to the and to the of J. R. J. Cell. Physiol. 86: PubMed Scopus Google Scholar). The amebocytes were from plasma by at for at The was to a through a and LPS affinity was by LPS from E. with CNBr-activated Sepharose CL-4B to the of with the of 2 of gel by the molecular mass of LPS to be of protease hemolymph was through three affinity with Sepharose CL-4B, staphylococcal protein A-Sepharose CL-4B, and LPS-Sepharose CL-4B, The were with buffer and at the of with at column of the buffer containing 1 m until a was The were from TPL-1 from the Sepharose CL-4B, which as the affinity the column was eluted with the buffer containing m GlcNAc. the lipopolysaccharide-binding protein (TPL-2), the LPS column was eluted with the buffer containing m and KCl. was to the containing the proteins to The was by at for and in The was at performance liquid chromatography was on an system with a column a of for protein and with a column a of for The of A and were acid at and at and peptides were eluted from with of B. for proteins and peptides were at and purified by was and in containing m with and with three of were The protein was with at for The peptides were by as described above column was to for the of gel was with with by with and with acid as described Biochem. PubMed Scopus Google Scholar). were by the to a mass for were to and and in to a column m The column was at to a of 1 as was at 60 for 1 to in 1 and to in The were by on the system a of at and to at of from the and from were on an protein The from to 20 The were by the TPL-1 and proteins from the of Sepharose CL-4B and LPS-affinity were purified by to a and a as by (15Chiou S.T. Chen Y.W. Chen S.C. Chao C.F. Liu T.Y. J. Biol. Chem. 2000; 275: 1630-1634Abstract Full Text Full Text PDF PubMed Scopus (30) Google Scholar). Each of the purified protein was with complete and a by the Chen S.T. S.C. J. Immunol. Scopus (24) Google Scholar). were and for the were at were from plasma by staphylococcal protein A column were by and in the Western blot and The of the specific was by or were on and to an Amersham Pharmacia with of The were with in with 20 and with specific were with in the and developed by the Amersham Pharmacia were with EZ-Link as described by the The were with with for 1 at to block The gel was with the and with horseshoe crab plasma at with the the was in the for and the was to Western blot the of bacteria in a of and were and the was a of A. PubMed Scopus Google Scholar). The of bacteria in the was by the of the at of with a The of was is to The with the bacteria were with buffer 20 in and the were with in the A TPL-1 or TPL-2 in buffer in was to and for 2 at or were to and for 2 at The TPL-1 and TPL-2 were with bacteria E. coli or to with these was to and the were for 2 at with the of in buffer was to and at for The reaction was by the of of 2 and the at was m GlcNAc and m GlcNAc the binding of TPL-1 and to bacteria, these as for the binding of TPL-1 and TPL-2 was on a All were in acid at and in the were from an of T. tridentatus. the and hemocytes were and in liquid were from the and were purified with chromatography Pharmacia The cDNA was with a and and was The cDNA was as a in The were as primer from of sequence and primer from of primer from of sequence and as The of cDNA was in a with the for 2 for for for 1 for were on a and E. coli by the sequence reaction was the were to electrophoresis on an and The of cDNA of TPL-1 and TPL-2 were by the cDNA cloning system with gene-specific was and was to and for was and was to and The for of TPL-1 were as to and All the were to the The and cDNA for TPL-1 and cDNA for TPL-2 were and as described T. J. Molecular A Scholar). These were to was purified from as described T. J. Molecular A Scholar). The for were on amino acid and TPL-2 and K. T. T. Iwanaga S. Kawabata S. J. Biol. Chem. 1999; Full Text Full Text PDF Scopus Google and The reaction was the for 2 for for for 1 for for for 1 for for for 1 for The was and as described T. J. Molecular A Scholar). plasma of horseshoe crabs through affinity two lectins were purified and which binds to Sepharose and which binds to the LPS of E. The of Sepharose CL-4B as a to the of plasma through the CL-4B, the of TPL-1 from TPL-1 and TPL-2 could be eluted from their affinity column with 2 m or In the previous study, both proteins were eluted from the affinity column with buffer containing or 2 m (15Chiou S.T. Chen Y.W. Chen S.C. Chao C.F. Liu T.Y. J. Biol. Chem. 2000; 275: 1630-1634Abstract Full Text Full Text PDF PubMed Scopus (30) Google Scholar). eluted with or 2 m of the agents. In the present study, m GlcNAc was found in TPL-1 from the Sepharose CL-4B of TPL-2 from the LPS-affinity column 2 m in to m GlcNAc. eluted with GlcNAc in solution of GlcNAc and can be to the affinity of the plasma through the affinity all of the proteins that bind to Sepharose CL-4B and to the of the with of other proteins were eluted with m or The of the binding of TPL-1 to the Sepharose CL-4B and by GlcNAc is Sepharose is a form of of of and an Sepharose is from Sepharose by with in the of chain and of the of the other J. T. J. Scopus Google Scholar). TPL-1 binds to this in a was isolated from the of horseshoe crabs by chromatography and shown to high affinity for both GlcNAc and S. Kawabata S. T. T. Iwanaga S. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). was from the of horseshoe crab by an affinity column of bovine to Sepharose and eluted from the column with GlcNAc T. Kawabata S. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). the plasma of horseshoe tachylectin and were an and of the proteins by GlcNAc (14Gokudan S. Muta T. Tsuda R. Koori K. Kawahara T. Seki N. Mizunoe Y. Wai S.N. Iwanaga S. Kawabata S. Proc. Natl. Acad. Sci. U. S. A. 1999; 96: 10086-10091Crossref PubMed Scopus (189) Google Scholar). a common specificity of binding to moiety of and sequence with these proteins bind to Sepharose CL-4B and be eluted from by is TPL-2 can be eluted from the LPS-affinity column with buffer containing LPS and 2 m KCl. However, the TPL-2 could be from LPS. GlcNAc is a of the LPS in the of LPS-affinity to protein. TPL-1 and TPL-2 isolated for about and of the hemolymph Although the of TPL-1 the of TPL-2 the have been in 1 TPL-1 showed two major at a at two at and at kDa. protein at and were In Western blot the with these protein in 1 and 1 with molecular masses and were in both the and plasma 1 and 1 TPL-2 with serum 1 the specificity of the antibodies. (15Chiou S.T. Chen Y.W. Chen S.C. Chao C.F. Liu T.Y. J. Biol. Chem. 2000; 275: 1630-1634Abstract Full Text Full Text PDF PubMed Scopus (30) Google Scholar), we described a protein with a molecular mass of as the major protein eluted by m from Sepharose m GlcNAc solution as the protein was A and The protein (TPL-1) was shown to have sequence with the sequence of (15Chiou S.T. Chen Y.W. Chen S.C. Chao C.F. Liu T.Y. J. Biol. Chem. 2000; 275: 1630-1634Abstract Full Text Full Text PDF PubMed Scopus (30) Google Scholar). The sequence 232 amino acid residues for TPL-1 The deduced amino acid sequence of TPL-1 shares a 65% identity to T. Kawabata S. Iwanaga S. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar), identified in the large granules of amebocytes of the horseshoe crab, and identity to T. Kawabata S. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar), an lectin in of the horseshoe crab A is the of a N-glycosylation site in TPL-1 and in the other two intracellular proteins, and Although and sequence with they and biochemical in and affinity to other proteins T. Kawabata S. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). TPL-1 also a identity to and of whose function is as J. J. R. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar). A sequence showed TPL-1 and other proteins and the of TPL-1 with and of T. tridentatus and and of The was and of residues are shown in of the in TPL-1 are involved in on sequence and of positions with T. Kawabata S. Iwanaga S. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar), the molecular mass of TPL-1 is This with the TPL-1 found by mass The of TPL-1 is a protein. showed other major with a molecular mass of and of and The mass of the TPL-1 and the TPL-1 can be to the of two and three on the The and the TPL-1 of TPL-1 as (15Chiou S.T. Chen Y.W. Chen S.C. Chao C.F. Liu T.Y. J. Biol. Chem. 2000; 275: 1630-1634Abstract Full Text Full Text PDF PubMed Scopus (30) Google Scholar). The Western blot and the mass the of and The to a of the the purified TPL-2 showed major protein with a mass of about 36 and a of about both 1 and 1 In Western blot these protein with the TPL-2 1 The plasma and through the affinity showed protein of and molecular with serum in the 1 and of a protein in the TPL-1 with serum 1 the specificity of antibodies. The deduced amino acid sequence of TPL-2 showed a identity with of the positions to Although a site, is present in this site is in Tachylectin-3 is a nonglycosylated intracellular protein isolated from the large of the of deduced amino acid of TPL-2 related found in T. tridentatus. The was of residues are shown in In previous report (15Chiou S.T. Chen Y.W. Chen S.C. Chao C.F. Liu T.Y. J. Biol. Chem. 2000; 275: 1630-1634Abstract Full Text Full Text PDF PubMed Scopus (30) Google Scholar), the residues and were as was shown as and as of the GlcNAc eluted TPL-2 showed in of acid in and of acid as T. Kawabata S. Iwanaga S. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google sequence as and both residues and as The sequence shown above and the sequence of TPL-2 (15Chiou S.T. Chen Y.W. Chen S.C. Chao C.F. Liu T.Y. J. Biol. Chem. 2000; 275: 1630-1634Abstract Full Text Full Text PDF PubMed Scopus (30) Google Scholar). The of acid at the that site of TPL-2 at this The sequence 128 amino acid residues for of the in TPL-2 are in on sequence and of positions with K. T. T. Iwanaga S. Kawabata S. J. Biol. Chem. 1999; Full Text Full Text PDF Scopus Google Scholar), the molecular mass of TPL-2 be showed a major with a mass of and two with masses of and the of the by mass and the mass of could be to a of of about 20 can be The of in TPL-2 was by and by of the TPL-2 eluted by GlcNAc. The of in this study is similar to the (15Chiou S.T. Chen Y.W. Chen S.C. Chao C.F. Liu T.Y. J. Biol. Chem. 2000; 275: 1630-1634Abstract Full Text Full Text PDF PubMed Scopus (30) Google Scholar). the of the be to the of to the molecular mass of a the of and as and This to a of 20 with a molecular mass of which is to the of the by mass and the mass of from the deduced amino acid sequence of the The of Western blot and mass of TPL-2 that the the the the and the the of TPL-2 contains with a that could form TPL-2 purified by affinity column exists as a dimer and 1 In the plasma, TPL-2 and exist as oligomers of molecular masses 1 and on sequence TPL-2 and tachylectin-3, was to the of for the were found to for proteins with similar amino acid sequence to and The that a homologous family of for TPL-2 and TPL-2 binds to LPS from E. This binding was the for the LPS-affinity The binding of TPL-2 to LPS is of since or was to TPL-2 from the LPS-affinity In this TPL-2 from the protein (8Minetti C.S.A. Lin Y. Cislo T. Liu T.-Y. J. Biol. Chem. 1991; 266: 20773-20780Abstract Full Text PDF PubMed Google Scholar) and T. Kawabata S. Iwanaga S. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). the proteins, the site of with LPS has been sequence were among a of these proteins J. J. R. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar). TPL-2 with other proteins, the protein purified by a LPS-affinity chromatography (8Minetti C.S.A. Lin Y. Cislo T. Liu T.-Y. J. Biol. Chem. 1991; 266: 20773-20780Abstract Full Text PDF PubMed Google Scholar), and isolated in this study, from most other proteins with a of a The of these proteins has been to be an in their with the LPS Science. 1990; PubMed Scopus Google Scholar). It could be that the of the amino acid in the of the protein is the of the protein for the binding to LPS. In this is that are three of amino acids in the TPL-2 sequence that be for binding to LPS. In to TPL-1 and TPL-2 to form with other with molecular masses of about 1 The of this in the two 1 1 that the in the The with molecular mass of about 1 and most likely from the to the in TPL-1 were from the gel and with in Western Although the by which and of TPL-1 and TPL-2 are to be the physiological function of TPL-1 and TPL-2 could be related to their to form of interlocking molecules to immobilize and entrap the invading TPL-1 and TPL-2 have been shown to bind to three of bacteria, S. coli in a and The specificity of the binding is by with 2 m Although both and plasma proteins bind to bacteria, plasma proteins bind to bacteria the of purified in binding to binding site of TPL-1 and TPL-2 with these bacteria be at the NAc moiety of the GlcNAc-MurNAc peptidoglycan. lectins with affinity for the S. Kawabata S. T. T. Iwanaga S. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar), T. Kawabata S. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar), and and (14Gokudan S. Muta T. Tsuda R. Koori K. Kawahara T. Seki N. Mizunoe Y. Wai S.N. Iwanaga S. Kawabata S. Proc. Natl. Acad. Sci. U. S. A. 1999; 96: 10086-10091Crossref PubMed Scopus (189) Google Scholar), could as innate defense molecules by binding to the of In to adaptive immune system in of specific antigen receptors and the innate immune system uses germline-encoded receptors for recognition of common antigens on the surface of microbial such as proteases (4Medzhitov R. Janeway Jr., C.A. Cell. 1997; 91: 295-298Abstract Full Text Full Text PDF PubMed Scopus (1965) Google Scholar), J.A. Reichhart J.M. Curr. Opin. Immunol. 1996; 8: PubMed Scopus Google Scholar), lectins R. Janeway Jr., C.A. Curr. Opin. Immunol. 1998; 10: PubMed Scopus Google Scholar), and peptides (5Muta T. Iwanaga S. Curr. Opin. Immunol. 1996; 8: 41-47Crossref PubMed Scopus (228) Google Scholar, 6Iwanaga S. Kawabata S. Muta T. J. Biochem. ( Tokyo ). 1998; 123: 1-15Crossref PubMed Scopus (253) Google Scholar). They in concert for the and of the invading pathogens. The lectins in the hemolymph are to and and to as the of defense the of pathogens. The lectins and in the of other defense which are for the and of the invading pathogens.