Purification and characterization of human serum C-reactive protein.
A simple and rapid purification method for human serum C-reactive protein (CRP) was developed. CRP was strongly adsorbed on a DEAE-cellulose column and was easily separated from other serum proteins. CRP was purified approximately 1,000-fold with a high yield (50%). The final preparation showed a single band as judged by SDS-polyacrylamide gel electrophoresis, polyacrylamide gel disc electrophoresis, and polyacrylamide gel isoelectric focusing. The fluorescence of the complex of CRP and 8-anilino-1-naphthalene sulfonate (ANS) changed with change of the pH, suggesting that CRP may show pH-dependent conformational change. This finding could account for the peculiar behavior of the protein in isoelectric focusing; it shows an isoelectric point of 7.4 when the starting pH is 7.0, whereas it shows two isoelectric points, 5.3 and 7.4, when the starting pH is 5.5. Ca2+-dependent change of the fluorescence of the complex of CRP and ANS was also detected. These results suggest a pH- and Ca2+-dependent conformational change of CRP.
D000596:Amino Acids / Q000032:analysis; D002097:C-Reactive Protein / Q000097:blood* / Q000302:isolation & purification; D002118:Calcium; D055598:Chemical Phenomena; D002621:Chemistry; D002848:Chromatography, DEAE-Cellulose; D004589:Electrophoresis, Disc; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D013050:Spectrometry, Fluorescence; D013535:Suspensions
D000596:Amino Acids; D013535:Suspensions; D002097:C-Reactive Protein; D002118:Calcium