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P1 1:30 Crustacean immunity
Abstracts of the 7th Congress of the ISDCI: Session P PI
205
1:30
CRUSTACEAN
IMMUNITY
Kenneth Siiderhill Department of Physiological Botany,University of Uppsala,Villavagen 6,752 36 UppsalaSweden Crustaceaens like most other arthropods have defence systems which can recognise and respond to pathogens. The prophenoloxidase activating system has heen shown to he ahle to fullfil such a function and the structural and functional properties of different components of this system will be reviewed. One of these molecules is pacifastin which is an efficient inhihitor of the prophenoloxidase activating system (ppA) and which consists of 9 inhihitor subunits and 3 transferrin chains linked together by covalent linkages with a mass of 155 kD.Another proPO-protein which recently has been cloned is the ppA and it was found to be a proenzyme and activated by cleavage resulting in one Nterminal (190 aa) and one C-terminal (232 aa) domain and the latter contains the serine proteinase domain. Since many immune proteins in crayfish have heen shown to he redox enzymes containing iron or cupper in their active sites we have also hegun to study proteins involved in iron hemostasis and to date chamcterised a numher of such proteins some of which will he discussed.
P2 2:Oo
CDNA CLONING OF THE CLOTTING PROTEIN FROM THE FRESHWATER CRAYFISH PACZFASTACUS LENZUSCULUS Ruigong Wang, Martin Hall, Kenneth Stklerhtil, Department of Physiological Botany, Uppsala University, VillavLgen 6, S-752 36 Uppsala, Sweden
The clotting protein (CP) is responsihle for the clot formation in the hemolymph of the fpshwater crayfish Puci$ustacusleniusculus. In crayfish, the soluble plasma CP is crosslinked together hy an endogenous transglutaminase from the crayfish hlood cell to form a very stahle and insoluble clot at the wound site. The N-terminal amino acid sequence and the internal peptide sequenceswere determined hy peptide sequencing. The 6,036hp of the cDNA of CP include a 159hp 5’-untranslated region, a 5,169hp open reading frame encoding 1,723 amino acid residues, which contains a hydrophobic signal peptide that consists of 15 amino acids, and a 708hp $-untranslated mgion. The authenticity of the cDNA sequence of CP is confirmed hy 17 peptide sequencescovering 3 18 amino acid residues of the ORF. The calculated molecular weight of the mature CP is 193.3 kDa, which agrees well with that of the purified CP suhunit (210 kDa) estimated hy SDS/PAGE. The overall deduced amino acid sequence of CP is similar to vitellogenins, particularly to insect vitellogenins, a female-specific protein important for development. A sequence hefore the cleavage site of endoproteases, RXRR, is highly eonserved in tie insect vitellognm family, is also found in the appropriate position in CP. The locations of cysteines are relatively conserved in CP and the insect vitellogenins, particularly at the C-terminus of the proteins. However, there is no set-me-richcluster in CP, which is usually found in vitellogenins. Northern analysis indicates that CP is synthesized in hepatopancreas in both male and female crayfish, and the size of CP mRNA is approximately 7kh. CP is also present and was purified from female crayfish. The N-terminal amino acid sequenceof the female CP is identical to that of the male CP.
205
1:30
CRUSTACEAN
IMMUNITY
Kenneth Siiderhill Department of Physiological Botany,University of Uppsala,Villavagen 6,752 36 UppsalaSweden Crustaceaens like most other arthropods have defence systems which can recognise and respond to pathogens. The prophenoloxidase activating system has heen shown to he ahle to fullfil such a function and the structural and functional properties of different components of this system will be reviewed. One of these molecules is pacifastin which is an efficient inhihitor of the prophenoloxidase activating system (ppA) and which consists of 9 inhihitor subunits and 3 transferrin chains linked together by covalent linkages with a mass of 155 kD.Another proPO-protein which recently has been cloned is the ppA and it was found to be a proenzyme and activated by cleavage resulting in one Nterminal (190 aa) and one C-terminal (232 aa) domain and the latter contains the serine proteinase domain. Since many immune proteins in crayfish have heen shown to he redox enzymes containing iron or cupper in their active sites we have also hegun to study proteins involved in iron hemostasis and to date chamcterised a numher of such proteins some of which will he discussed.
P2 2:Oo
CDNA CLONING OF THE CLOTTING PROTEIN FROM THE FRESHWATER CRAYFISH PACZFASTACUS LENZUSCULUS Ruigong Wang, Martin Hall, Kenneth Stklerhtil, Department of Physiological Botany, Uppsala University, VillavLgen 6, S-752 36 Uppsala, Sweden
The clotting protein (CP) is responsihle for the clot formation in the hemolymph of the fpshwater crayfish Puci$ustacusleniusculus. In crayfish, the soluble plasma CP is crosslinked together hy an endogenous transglutaminase from the crayfish hlood cell to form a very stahle and insoluble clot at the wound site. The N-terminal amino acid sequence and the internal peptide sequenceswere determined hy peptide sequencing. The 6,036hp of the cDNA of CP include a 159hp 5’-untranslated region, a 5,169hp open reading frame encoding 1,723 amino acid residues, which contains a hydrophobic signal peptide that consists of 15 amino acids, and a 708hp $-untranslated mgion. The authenticity of the cDNA sequence of CP is confirmed hy 17 peptide sequencescovering 3 18 amino acid residues of the ORF. The calculated molecular weight of the mature CP is 193.3 kDa, which agrees well with that of the purified CP suhunit (210 kDa) estimated hy SDS/PAGE. The overall deduced amino acid sequence of CP is similar to vitellogenins, particularly to insect vitellogenins, a female-specific protein important for development. A sequence hefore the cleavage site of endoproteases, RXRR, is highly eonserved in tie insect vitellognm family, is also found in the appropriate position in CP. The locations of cysteines are relatively conserved in CP and the insect vitellogenins, particularly at the C-terminus of the proteins. However, there is no set-me-richcluster in CP, which is usually found in vitellogenins. Northern analysis indicates that CP is synthesized in hepatopancreas in both male and female crayfish, and the size of CP mRNA is approximately 7kh. CP is also present and was purified from female crayfish. The N-terminal amino acid sequenceof the female CP is identical to that of the male CP.