Isolation and characterization of C-reactive protein and serum amyloid P component from bovine serum.
C-reactive protein (CRP) and serum amyloid P component (SAP), which are known as acute phase reactants in human and many other animals, were purified from cow sera. Affinity chromatography using HE agarose gel was the most effective method to isolate both CRP and SAP from a large volume of bovine serum. Separation of CRP and SAP from the mixed preparation could be performed by DEAE-cellulose column chromatography, gel permeation HPLC using TSK-G3000SW or affinity chromatography using phosphorylcholine derivatives of bovine serum albumin-conjugated Toyopearl HW 65. Bovine CRP and SAP were identified as genuine CRP- and SAP-class proteins by their cross reactivities with anti-human CRP and anti-human SAP, respectively, and by their homology in amino acid compositions compared with those of human CRP and SAP, respectively. Bovine CRP moved slower than beta-globulin, and bovine SAP moved in the beta-globulin region in agarose gel electrophoresis. Both of them gave single bands in native polyacrylamide gel electrophoresis (PAGE). Bovine CRP and SAP molecular weights were estimated to be 100,600 and 109,500 daltons respectively, by sedimentation equilibrium analysis. Bovine CRP showed 23K dalton subunits by sodium laurylsulfate-PAGE and bovine SAP showed 28K and 32K dalton subunits, both of which were glycosylated and had identical amino acid compositions, indicating that both CRP and SAP molecules are pentamers. In fact, they appeared to have pentameric disk-like configurations in electronmicroscopical examination.
Morimatsu
M
M
Sakai
H
H
Yoshimatsu
K
K
Minowa
O
O
Yamamoto
S
S
Yatomi
K
K
Fujinaga
T
T
Naiki
M
M
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002417:Cattle / Q000097:blood*; D002845:Chromatography; D004586:Electrophoresis; D008970:Molecular Weight; D011486:Protein C / Q000032:analysis / Q000302:isolation & purification*; D000683:Serum Amyloid P-Component / Q000032:analysis / Q000302:isolation & purification*
D000596:Amino Acids; D011486:Protein C; D000683:Serum Amyloid P-Component