Investigation of C-reactive protein binding to phosphatidyl choline by CZE and ESI-mass analysis.
A simple CZE has been developed to investigate the binding reactions between C-reactive protein (CRP) and native as well as Cu(2+) oxidized phosphatidyl choline (PC) in this study. An aqueous-organic solvent system containing 20% sodium phosphate, 70% methanol and 10% acetonitrile was selected as the optimal CZE separation buffer. Native PC showed a broad, higher mobility major peak and a sharp, lower mobility minor peak. Oxidized PC (ox-PC) appeared as two adjacent peaks. PC bound to CRP in the presence of metal ions Ca(2+) and Mg(2+). The absorbance at 214 nm for PC+CRP was about five times of that for PC. Surprisingly, ox-PC interacted much more weakly with CRP. The absorbance at 214 nm for ox-PC+CRP was close to that of ox-PC. Meanwhile, ESI-Mass analysis also suggested that four PC molecular species (C16:0/C18:1, C16:0/C18:2, C18:0/C18:2, C18:0/C18:1) bound with CRP since the ion intensities of PC+CRP were lower than that of PC alone. This study uses both CZE and ESI-Mass analysis to characterize the interaction between CRP and native as well as Cu(2+) oxidized PC, and provides new analytical methods for determining the binding reactions. In addition, it also gives new insight into the molecular interactions between CRP and native as well as oxidized PC.
Department of Chemistry, National Changhua University of Education, Changhua, Taiwan.
Yu-Non
YN
Peng
Yu-Ling
YL
Ho
Chung-Yu
CY
Wu
Mine-Yine
MY
Liu
D000097:Acetonitriles / Q000737:chemistry; D000818:Animals; D002097:C-Reactive Protein / Q000737:chemistry / Q000378:metabolism*; D019075:Electrophoresis, Capillary / Q000379:methods*; D006801:Humans; D000432:Methanol / Q000737:chemistry; D010710:Phosphates / Q000737:chemistry; D010713:Phosphatidylcholines / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
D000097:Acetonitriles; D010710:Phosphates; D010713:Phosphatidylcholines; D002097:C-Reactive Protein; C018279:sodium phosphate; D000432:Methanol; C032159:acetonitrile