title: Identification of the binding site for plasma prekallikrein in human high molecular weight kininogen. A region from residues 185 to 224 of the kininogen light chain retains full binding activity.
abstract: We studied the ability of fragments of the light chain of human high molecular weight kininogen to bind to plasma prekallikrein. In a competitive fluorescence polarization assay, kallikrein-cleaved light chain (light chain-2; residues 49-255), a cyanogen bromide fragment (residues 185-242), and a tryptic peptide (T-7; residues 185-224) had binding affinities of approximately 20 nM, equivalent to the value for the intact light chain (residues 1-255) of high-molecular-weight kininogen. In contrast, fragments consisting of residues 49-184 and 243-255 showed no binding activity (Kd much greater than 1,000 nM). Direct titrations of fluorescein-labeled derivatives of light chain-2 and peptide T-7 with prekallikrein confirmed that T-7 retained full binding activity for prekallikrein (Kd = 12 +/- 2 nM for labeled light chain-2; Kd = 7 +/- 1 nM for labeled T-7). These results localize the binding site of high molecular weight kininogen for prekallikrein within a region of 40 amino acids (residues 185-224) that resides in the near carboxyl terminus of the light chain of kininogen.
mesh_terms: D001665:Binding Sites; D003488:Cyanogen Bromide; D016650:Fluorescein-5-isothiocyanate; D005452:Fluoresceins; D005454:Fluorescence Polarization; D005456:Fluorescent Dyes; D006801:Humans; D007610:Kallikreins / Q000502:physiology*; D007704:Kininogens / Q000097:blood*; D008970:Molecular Weight; D010446:Peptide Fragments / Q000097:blood; D011288:Prekallikrein / Q000502:physiology*; D013861:Thiocyanates; D014357:Trypsin
chemical_list: D005452:Fluoresceins; D005456:Fluorescent Dyes; D007704:Kininogens; D010446:Peptide Fragments; D013861:Thiocyanates; D011288:Prekallikrein; D007610:Kallikreins; D014357:Trypsin; D016650:Fluorescein-5-isothiocyanate; D003488:Cyanogen Bromide