Platelet thrombospondin interactions with human high and low molecular weight kininogens.
Multifunctional proteins, e.g. high molecular weight kininogen (HK, 120 kDa) and the homotrimer, thrombospondin (TSP, 540 kDa), which have more than one domain on a single polypeptide chain, are particularly well-suited to be structural elements of extracellular matrices because of their ability to bind to several macromolecules. We now demonstrate that 125I-high molecular weight kininogen (HKa) cleaved by purified kallikrein forms a complex with purified intact platelet TSP (540 kDa). HK also complexed with a proteolytic fragment (450 kDa) of TSP, lacking its three identical heparin-binding domains (HBD, 30 kDa), but failed to bind to a more extensively proteolysed molecule (210 kDa) lacking the C-terminal globular domain indicating that the binding on TSP-450 kDa is confined to the C-terminus. The binding of HK to intact TSP and to its 450 kDa fragment was of high affinity (Kd = 17-52 nM), specific, concentration dependent and saturable. Furthermore, we found both forms of the light chain (LC) of HK (56 and 46 kDa) resulting from cleavage by plasma kallikrein bound to both intact TSP and HBD independent of the presence of calcium ions. However, neither the epitope recognized by monoclonal antibody (MAb) C11C1 on domain 5 nor the prekallikrein binding site on domain 6 are involved, suggesting that the intervening proline-rich region may be the site of interaction. The heavy chain (HC) of HK required ionized calcium to bind to intact TSP or its 450 kDa homotrimer fragment.(ABSTRACT TRUNCATED AT 250 WORDS)
Sol Sherry Thrombosis Research Center, Department of Pathology and Laboratory Medicine, Temple University School of Medicine, Philadelphia, PA 19140.
R A
RA
DeLa Cadena
E G
EG
Wyshock
S P
SP
Kunapuli
R L
RL
Schultze
M
M
Miller
D A
DA
Walz
R W
RW
Colman
D001792:Blood Platelets / Q000378:metabolism*; D002118:Calcium / Q000097:blood; D006801:Humans; D015151:Immunoblotting; D007457:Iodine Radioisotopes; D007704:Kininogens / Q000097:blood*; D008562:Membrane Glycoproteins / Q000097:blood*; D008970:Molecular Weight; D011485:Protein Binding; D011869:Radioligand Assay; D019699:Thrombospondins
D007457:Iodine Radioisotopes; D007704:Kininogens; D008562:Membrane Glycoproteins; D019699:Thrombospondins; D002118:Calcium