Identification and characterization of a major bovine serum tyrosine-O-sulfate-binding protein as a complement factor H.
A major tyrosine-O-sulfate (TyrS)-binding protein present in bovine serum was purified to electrophoretic homogeneity using a combination of TyrS-Affi-Gel 10 affinity chromatography, DEAE-Bio-Gel A ion-exchange chromatography, and hydroxylapatite chromatography. The purified TyrS-binding protein migrated as doublet protein bands with apparent molecular weights of ca. 160,000, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions. N-termini of the two forms of purified TyrS-binding protein contain most likely identical sequence for the first fifteen amino acids residues, which displays a high degree of homology to those of human and mouse complement factor H. Furthermore, the purified TyrS-binding protein exhibited immunologic cross-reactivity with anti-human complement factor H. These results indicate the identity of the purified TyrS-binding protein being bovine complement factor H. The two forms of the purified bovine factor H were investigated with respect to the sensitivity to limited trypsin digestion. The high-molecular weight form was cleaved into two fragments with apparent molecular masses of, respectively, 45 kD and 125 kD. The low-molecular weight form was cleaved in a different manner to generate three major fragments with molecular masses of 25 kD, 45 kD and 100 kD, respectively. Limited V8 protease mapping of the two forms yielded similar, yet unidentical, peptide band patterns. Purified bovine factor H appeared to bind agarose-bonded heparin through its anion-binding domain and the binding was inhibited by the presence of free heparin or dextran sulfate.
Sakakibara
Y
Y
Department of Biochemistry, University of Texas Health Center at Tyler 75710.
Suiko
M
M
Fernando
P H
PH
Ohashi
T
T
Liu
M C
MC
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002352:Carrier Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D002417:Cattle; D017242:Complement Factor H / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D003429:Cross Reactions; D004247:DNA / Q000378:metabolism; D016264:Dextran Sulfate / Q000494:pharmacology; D006493:Heparin / Q000378:metabolism; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010449:Peptide Mapping; D000071228:Polyelectrolytes; D011108:Polymers / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D011487:Protein Conformation; D014443:Tyrosine / Q000031:analogs & derivatives / Q000378:metabolism
D000596:Amino Acids; D002352:Carrier Proteins; D017242:Complement Factor H; D004247:DNA; D016264:Dextran Sulfate; D006493:Heparin; D036341:Intercellular Signaling Peptides and Proteins; D000071228:Polyelectrolytes; D011108:Polymers; D014443:Tyrosine; C493495:CFH protein, human; C057501:tyrosine-O-sulfate-binding protein, Bos taurus; C005662:tyrosine O-sulfate