Induction of glycosylation in human C-reactive protein under different pathological conditions.
As an acute-phase protein, human C-reactive protein (CRP) is clinically important. CRPs were purified from several samples in six different pathological conditions, where their levels ranged from 22 to 342 microg/ml. Small, but significant, variations in electrophoretic mobilities on native PAGE suggested differences in molecular mass, charge and/or shape. Following separation by SDS/PAGE, they showed single subunits with some differences in their molecular masses ranging between 27 and 30.5 kDa, but for a particular disease, the mobility was the same for CRPs purified from multiple individuals or pooled sera. Isoelectric focusing (IEF) also indicated that the purified CRPs differed from each other. Glycosylation was demonstrated in these purified CRPs by Digoxigenin kits, neuraminidase treatment and binding with lectins. The presence of N-linked sugar moiety was confirmed by N-glycosidase F digestion. The presence of sialic acid, glucose, galactose and mannose has been demonstrated by gas liquid chromatography, mass spectroscopic and fluorimetric analysis. Matrix-assisted laser-desorption ionization analysis of the tryptic digests of three CRPs showed systematic absence of two peptide fragments, one at the N-terminus and the other near the C-terminus. Model-building suggested that the loss of these fragments exposed two potential glycosylation sites on a cleft floor keeping the protein-protein interactions in pentraxins and calcium-dependent phosphorylcholine-binding qualitatively unaffected. Thus we have convincingly demonstrated that human CRP is glycosylated in some pathological conditions.
Das
Tanusree
T
Immunobiology Division, Indian Institute of Chemical Biology, 4 Raja S C Mullick Road, Calcutta 700 032, India.
Sen
Asish K
AK
Kempf
Tore
T
Pramanik
Sumit R
SR
Mandal
Chhabinath
C
Mandal
Chitra
C
D000595:Amino Acid Sequence; D002097:C-Reactive Protein / Q000737:chemistry / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D005690:Galactose / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D007525:Isoelectric Focusing; D037102:Lectins / Q000378:metabolism; D008180:Lupus Erythematosus, Systemic / Q000378:metabolism / Q000473:pathology*; D008358:Mannose / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019158:N-Acetylneuraminic Acid / Q000378:metabolism; D012516:Osteosarcoma / Q000378:metabolism / Q000473:pathology*; D010767:Phosphorylcholine / Q000378:metabolism; D054198:Precursor Cell Lymphoblastic Leukemia-Lymphoma / Q000378:metabolism / Q000473:pathology*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D037102:Lectins; D010767:Phosphorylcholine; D002097:C-Reactive Protein; D019158:N-Acetylneuraminic Acid; D005947:Glucose; D008358:Mannose; D005690:Galactose