Type IV collagen-binding proteins of neutrophils: possible involvement of L-selectin in the neutrophil binding to type IV collagen.
To isolate type IV collagen-binding proteins, 125I-labeled human-neutrophil extracts were chromatographed on a type IV collagen-Sepharose column. The affinity chromatography-separated fraction contained the four radioactive proteins with apparent molecular masses of 28, 49, 67, and 95 kD on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Western blot analysis indicated that the 95-kD proteins contained both L-selectin and nonspecific cross-reacting antigen 90 (NCA90), and that the 67-kD protein was the 67-kD elastin/laminin-binding protein (67BP). The data obtained with the type IV collagen-affinity chromatography and the immunoaffinity chromatographies using anti-L-selectin and anti-NCA90 monoclonal antibodies (MoAbs) have shown that L-selectin is closely associated with 67BP and the 49-kD protein, and that NCA90 is associated with 67BP, the 28-kD and 49-kD proteins. Among these binding proteins, sialic acid residues were contained in 67BP, L-selectin, and NCA90, but not in the 28-kD and 49-kD proteins. Sialidase treatment completely abolished both the binding affinity of the type IV collagen-binding proteins to type IV collagen and the neutrophil adherence to type IV collagen-coated plastic. Thus, the sialic acid residues of 67BP, L-selectin, and NCA90 seem to be important for the binding of neutrophils to type IV collagen. Furthermore, L-selectin IgG chimeric protein directly bound to type IV collagen-Sepharose column, and anti-L-selectin MoAb DREG56 inhibited the neutrophil adherence to type IV collagen-coated plastic by 51%. These observations suggest that L-selectin likely plays a role in the neutrophil binding to type IV collagen, although neutrophils have several kinds of adhesion molecules for type IV collagen such as L-selectin, NCA90, 67BP, and the 28-kD and 49-kD proteins.
Department of Biochemistry, School of Medicine, Juntendo University, Tokyo, Japan.
K
K
Iwabuchi
I
I
Nagaoka
A
A
Someya
T
T
Yamashita
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology; D000951:Antigens, Neoplasm; D015815:Cell Adhesion Molecules; D002846:Chromatography, Affinity; D003094:Collagen / Q000145:classification / Q000378:metabolism*; D006801:Humans; D016023:Integrins / Q000302:isolation & purification / Q000378:metabolism*; D019041:L-Selectin / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000302:isolation & purification / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D019158:N-Acetylneuraminic Acid; D009439:Neuraminidase / Q000494:pharmacology; D009504:Neutrophils / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects; D011956:Receptors, Cell Surface / Q000302:isolation & purification / Q000378:metabolism; D038981:Receptors, Collagen; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012794:Sialic Acids / Q000032:analysis
D000911:Antibodies, Monoclonal; D000951:Antigens, Neoplasm; D015815:Cell Adhesion Molecules; D016023:Integrins; D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; D038981:Receptors, Collagen; D011993:Recombinant Fusion Proteins; D012794:Sialic Acids; C054901:elastin-binding proteins; D019041:L-Selectin; D003094:Collagen; D009439:Neuraminidase; D019158:N-Acetylneuraminic Acid