title: Isolation and characterization of SAP and CRP, two pentraxins from Pangasianodon (Pangasius) hypophthalmus.
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 microg SAP and 56 microg 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 beta-pleated sheets are dominating the secondary structure. Upon removing the Ca(2+) 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.
authors.affiliation: Laboratory of Protein Chemistry, Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussel, Belgium.
authors.forename: Duong Thi
authors.initials: DT
authors.lastname: Huong Giang
authors.forename: Edilbert
authors.initials: E
authors.lastname: Van Driessche
authors.forename: Isabel
authors.initials: I
authors.lastname: Vandenberghe
authors.forename: Bart
authors.initials: B
authors.lastname: Devreese
authors.forename: Sonia
authors.initials: S
authors.lastname: Beeckmans
mesh_terms: D000371:Agglutination; D000595:Amino Acid Sequence; D000818:Animals; D001419:Bacteria / Q000378:metabolism; D015153:Blotting, Western; D002097:C-Reactive Protein / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D002397:Catfishes / Q000235:genetics* / Q000378:metabolism*; D002850:Chromatography, Gel; D003429:Cross Reactions; D004591:Electrophoresis, Polyacrylamide Gel; D004912:Erythrocytes / Q000378:metabolism; D007031:Hypothalamus / Q000378:metabolism*; D008969:Molecular Sequence Data; D011817:Rabbits; D016415:Sequence Alignment; D000683:Serum Amyloid P-Component / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D013057:Spectrum Analysis
chemical_list: D000683:Serum Amyloid P-Component; D002097:C-Reactive Protein