The structure of human thrombospondin, an adhesive glycoprotein with multiple calcium-binding sites and homologies with several different proteins

Thrombospondin is one of a class of adhesive glycoproteins that mediate cell-to-cell and cell-to-matrix interactions. We have used two monoclonal antibodies to isolate cDNA clones of thrombospondin from a human endothelial cell cDNA library and have determined the complete nucleotide sequence of the coding region. Three regions of known amino acid sequence of human platelet thrombospondin confirm that the clones are authentic. Three types of repeating amino acid sequence are present in thrombospondin. The first is 57 amino acids long and shows homology with circumsporozoite protein from Plasmodium falciparum. The second is 50-60 amino acids long and shows homology with epidermal growth factor precursor. The third occurs as a continuous eightfold repeat of a 38- residue sequence; structural homology with parvalbumin and calmodulin indicates that these repeats constitute the multiple calcium-binding sites of thrombospondin. The amino acid sequence arg-gly-asp-ala is included in the last type 3 repeat. This sequence is probably the site for the association of thrombospondin with cells. In addition, localized homologies with procollagen, fibronectin, and von Willebrand factor are present in one region of the thrombospondin molecule.

[1]  R. Hynes,et al.  Analysis of platelet adhesion with a radioactive chemical crosslinking reagent: Interaction of thrombospondin with fibronectin and collagen , 1982, Cell.

[2]  P. McKeown-Longo,et al.  Cultured human fibroblasts synthesize and secrete thrombospondin and incorporate it into extracellular matrix. , 1983, Proceedings of the National Academy of Sciences of the United States of America.

[3]  A. Kornblihtt,et al.  Primary structure of human fibronectin: differential splicing may generate at least 10 polypeptides from a single gene. , 1985, The EMBO journal.

[4]  C. Dang,et al.  Localization of a fibrinogen calcium binding site between gamma-subunit positions 311 and 336 by terbium fluorescence. , 1985, The Journal of biological chemistry.

[5]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[6]  David D. Roberts,et al.  Thrombospondin binds falciparum malaria parasitized erythrocytes and may mediate cytoadherence , 1985, Nature.

[7]  J. Lawler,et al.  Thrombospondin interactions with fibronectin and fibrinogen. Mutual inhibition in binding. , 1984, European journal of biochemistry.

[8]  J. Coligan,et al.  Isolation and characterization of a high molecular weight glycoprotein from human blood platelets. , 1978, The Journal of biological chemistry.

[9]  R. Silverstein,et al.  Complex formation of platelet thrombospondin with plasminogen. Modulation of activation by tissue activator. , 1984, The Journal of clinical investigation.

[10]  R. Hynes,et al.  Molecular biology of fibronectin. , 1985, Annual review of cell biology.

[11]  V. Dixit,et al.  Mapping of epitopes for monoclonal antibodies against human platelet thrombospondin with electron microscopy and high sensitivity amino acid sequencing , 1985, The Journal of cell biology.

[12]  J. Lawler,et al.  The release of heparin binding peptides from platelet thrombospondin by proteolytic action of thrombin, plasmin and trypsin. , 1981, Thrombosis research.

[13]  V. Dixit,et al.  Monoclonal antibodies that recognize calcium-dependent structures of human thrombospondin. Characterization and mapping of their epitopes. , 1986, The Journal of biological chemistry.

[14]  J. Turk,et al.  Thiol-disulfide exchange by thrombospondin: evidence for a thiol and a disulfide bond protected by calcium. , 1986, Archives of biochemistry and biophysics.

[15]  J. Connolly,et al.  Thrombin and Chymotrypsin Interactions with Thrombospondin a , 1986, Annals of the New York Academy of Sciences.

[16]  F. Sanger,et al.  DNA sequencing with chain-terminating inhibitors. , 1977, Proceedings of the National Academy of Sciences of the United States of America.

[17]  D. Lipman,et al.  Rapid and sensitive protein similarity searches. , 1985, Science.

[18]  S. Latt,et al.  Human von Willebrand factor (vWF): isolation of complementary DNA (cDNA) clones and chromosomal localization. , 1985, Science.

[19]  J. Myers,et al.  Human proα1(I) collagen gene structure reveals evolutionary conservation of a pattern of introns and exons , 1984, Nature.

[20]  D. Mosher,et al.  Localization of thrombospondin in clots formed in situ. , 1985, Blood.

[21]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[22]  M. Kozak.  Point mutations define a sequence flanking the AUG initiator codon that modulates translation by eukaryotic ribosomes , 1986, Cell.

[23]  D. Walz,et al.  Antibodies against a 23Kd heparin binding fragment of thrombospondin inhibit platelet aggregation. , 1984, Biochemical and biophysical research communications.

[24]  J. Lawler,et al.  Physical characterization of platelet thrombospondin. , 1981, The Journal of biological chemistry.

[25]  J. Coligan,et al.  Structure of thrombospondin. , 1984, The Journal of biological chemistry.

[26]  C. Southan,et al.  COVALENT STRUCTURE OF FIBRINOGEN , 1983, Annals of the New York Academy of Sciences.

[27]  P. McKeown-Longo,et al.  Binding and degradation of platelet thrombospondin by cultured fibroblasts , 1984, The Journal of cell biology.

[28]  A. Nurden,et al.  Immunocytochemical study of the binding of fibrinogen and thrombospondin to ADP- and thrombin-stimulated human platelets. , 1985, Blood.

[29]  E. Ruoslahti,et al.  Arg-Gly-Asp: A versatile cell recognition signal , 1986, Cell.

[30]  M. Ginsberg,et al.  Interaction of thrombospondin with resting and stimulated human platelets. , 1986, The Journal of biological chemistry.

[31]  J. Lawler.  The structural and functional properties of thrombospondin. , 1986, Blood.

[32]  J. Lawler,et al.  Evidence for calcium-sensitive structure in platelet thrombospondin. Isolation and partial characterization of thrombospondin in the presence of calcium. , 1982, The Journal of biological chemistry.

[33]  R. Hynes,et al.  Isolation and analysis of cDNA and genomic clones of fibronectin and its receptor. , 1987, Methods in enzymology.

[34]  E Ruoslahti,et al.  Platelet membrane glycoprotein IIb/IIIa: member of a family of Arg-Gly-Asp--specific adhesion receptors. , 1986, Science.

[35]  W. Rutter,et al.  Structure of a mouse submaxillary messenger RNA encoding epidermal growth factor and seven related proteins. , 1983, Science.

[36]  A. Ullrich,et al.  Nucleotide sequence of epidermal growth factor cDNA predicts a 128,000-molecular weight protein precursor , 1983, Nature.

[37]  P. Bornstein,et al.  Platelet-derived growth factor and heparin-like glycosaminoglycans regulate thrombospondin synthesis and deposition in the matrix by smooth muscle cells , 1985, The Journal of cell biology.

[38]  S. C. Hubbard,et al.  Synthesis and processing of asparagine-linked oligosaccharides. , 1981, Annual review of biochemistry.

[39]  R. Hynes,et al.  Interaction of fibronectin with its receptor on platelets , 1985, Cell.

[40]  V. Nussenzweig,et al.  Circumsporozoite proteins of malaria parasites , 1985, Cell.

[41]  P. Bornstein,et al.  Thrombospondin: synthesis and secretion by cells in culture , 1982, The Journal of cell biology.

[42]  R. Doolittle.  Similar amino acid sequences: chance or common ancestry? , 1981, Science.

[43]  S. Henikoff.  Unidirectional digestion with exonuclease III creates targeted breakpoints for DNA sequencing. , 1984, Gene.

[44]  R. Hardison,et al.  The isolation of structural genes from libraries of eucaryotic DNA , 1978, Cell.

[45]  R. Doolittle,et al.  - cross-linking sites in human and bovine fibrin. , 1971, Biochemistry.

[46]  B. Vogelstein,et al.  Preparative and analytical purification of DNA from agarose. , 1979, Proceedings of the National Academy of Sciences of the United States of America.

[47]  J. Lawler,et al.  Cooperative binding of calcium to thrombospondin. The effect of calcium on the circular dichroism and limited tryptic digestion of thrombospondin. , 1983, The Journal of biological chemistry.

[48]  P. Y. Chou,et al.  Prediction of the secondary structure of proteins from their amino acid sequence. , 2006 .

[49]  V. Dixit,et al.  Isolation and characterization of a heparin-binding domain from the amino terminus of platelet thrombospondin. , 1984, The Journal of biological chemistry.

[50]  R. Kretsinger.  CRYSTALLOGRAPHIC STUDIES OF CALMODULIN AND HOMOLOGS , 1980, Annals of the New York Academy of Sciences.

[51]  Prediction of the secondary structure of platelet factor 4 and beta-thromboglobulin from their amino acid sequences. , 1981, Thrombosis research.

[52]  R. Hynes,et al.  Three different fibronectin mRNAs arise by alternative splicing within the coding region , 1983, Cell.

[53]  Ronald W. Davis,et al.  Immunoscreening λgtll Recombinant DNA Expression Libraries , 1985 .

[54]  J. Weber,et al.  Structure of the gene encoding the immunodominant surface antigen on the sporozoite of the human malaria parasite Plasmodium falciparum. , 1984, Science.

[55]  L. Leung,et al.  Complex formation of platelet thrombospondin with fibrinogen. , 1982, The Journal of clinical investigation.

[56]  P. Vischer,et al.  Synthesis, intracellular processing and secretion of thrombospondin in human endothelial cells. , 1985, European journal of biochemistry.

[57]  C. Lapière,et al.  Amino acid sequence of the aminoterminal segment of dermatosparactic calf-skin procollagen type I. , 1979, European journal of biochemistry.

[58]  L. Westrick,et al.  Incorporation of thrombospondin into fibrin clots. , 1985, The Journal of biological chemistry.

[59]  P. Bornstein,et al.  Location and partial characterization of the heparin-binding fragment of platelet thrombospondin. , 1984, Thrombosis research.

[60]  J. Connolly,et al.  The structure of human platelet thrombospondin. , 1985, The Journal of biological chemistry.

[61]  V. Dixit,et al.  A monoclonal antibody against human thrombospondin inhibits platelet aggregation. , 1985, Proceedings of the National Academy of Sciences of the United States of America.

[62]  J. Mcdonagh,et al.  Amino acid sequence of the factor XIIIa acceptor site in bovine plasma fibronectin , 1981, FEBS letters.

[63]  V. Nussenzweig,et al.  Structure of the plasmodium knowlesi gene coding for the circumsporozoite protein , 1983, Cell.

[64]  P. Bornstein,et al.  Interactions of thrombospondin with extracellular matrix proteins: selective binding to type V collagen , 1984, The Journal of cell biology.