Isolation and characterization of SAP and CRP,two pentraxins from Pangasianodon (Pangasius) hypophthalmus |
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| Authors: | Duong Thi Huong Giang Edilbert Van Driessche Isabel Vandenberghe Bart Devreese Sonia Beeckmans |
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| Institution: | 1. International University, VNU HCMC, Quarter 6, Linh Trung Ward, Thu Duc District, Ho Chi Minh City, Viet Nam;2. Institute for Future Environment, Queensland University of Technology, GPO Box 2434, Brisbane, QLD 4001, Australia;3. Science and Engineering Faculty, Queensland University of Technology, GPO Box 2434, Brisbane, QLD 4001, Australia;4. CSIRO Livestock Industries, Queensland Biosciences Precinct, QLD 4057, Australia;1. Carbohydrate and Protein Biotechnology Laboratory, Department of Biotechnology, Punjabi University, Patiala, 147 002, Punjab, India;2. Nanomedicine-Laboratory of Immunology and Molecular Biomedical Research (NLIMBR), School of Medicine (SoM), Centre for Molecular and Medical Research (C-MMR), Faculty of Health, Deakin University, Geelong, Victoria 3217, Australia;1. Department of Molecular and Cellular Biology, University of California, Davis, CA, USA;2. Marine Biological Laboratory, Woods Hole, MA, USA |
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| Abstract: | From the serum of Pangasianodon hypophthalmus, two proteins were isolated by affinity chromatography on Sepharose and phosphorylcholine–Sepharose. Their binding on the affinity matrices critically depends on the presence of Ca2+ ions. N-terminal sequencing and sequencing of internal tryptic peptides identified the proteins as pentraxins and from their binding properties they are identified as SAP (serum amyloid P component) and CRP (C-reactive protein). Per ml serum, 36 μg SAP and 56 μg CRP was purified. Upon gel filtration, both the SAP and CRP elute as trimers of respectively 24 kDa and 28 kDa subunits. Both proteins are devoid of inter-chain disulfide bonds. Both SAP and CRP are glycosylated and agglutinate rabbit erythrocytes and pathogenic bacteria Edwardsiella ictaluri and Aeromonas hydrophila, but not Micrococcus lysodeikticus or Escherichia coli. Haemagglutination of SAP and CRP is inhibited by galactose (MIC = 1 mM) and by phosphorylcholine (MIC = 1–2 mM), respectively. Circular dichroism studies revealed that antiparallel β-pleated sheets are dominating the secondary structure. Upon removing the Ca2+ ions by EDTA, slight structural changes are observed by CD spectroscopy in the near-UV region. Immunodiffusion shows that P. hypophthalmus SAP and CRP do not cross-react. |
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