Direct evidence for multifacial contacts between high molecular weight kininogen and plasma prekallikrein.
HK31 (S565-K595) has previously been shown to encompass the binding domain for plasma prekallikrein (PK) within domain 6 of high molecular weight kininogen (HK). The complementary binding domain for HK within PK is mapped to PK56 (F56-G86), in the apple 1 domain, and to PK266 (K266-C295), in the apple 4 domain. Isothermal titration calorimetry was used to directly monitor binding among HK31, PK56, and PK266. Either PK peptide binds to HK31 in 1:1 stoichiometry, regardless of whether a binary complex is first formed between PK266 and HK31 or between PK56 and HK31. Binding of the alternate PK peptide into a ternary complex is facilitated nearly 2-fold. The ternary complex consists of 1:1:1 HK31:PK56:PK266. Furthermore, binary and ternary complex formation is entropically driven and thermodynamically favored, suggesting that the conformational changes accompany binding. Fluorescence emission spectroscopy revealed that binding of PK56 caused a limited decrease in intrinsic tryptophan fluorescence emission intensity of HK31 while binding of PK266 to HK31 or the complex of HK31/PK56 had no such effect. We conclude that the two PK peptides bind to the HK peptide at different sites. The binding between HK and PK is likely due to conformational changes which serve to juxtapose the PK binding domain within HK with the HK binding site involving two spatial proximity segments.
Lin
Y
Y
Sol Sherry Thrombosis Research Center, Temple University School of Medicine, Philadelphia, Pennsylvania 19140, USA.
Shenoy
S S
SS
Harris
R B
RB
Colman
R W
RW
D000595:Amino Acid Sequence; D001665:Binding Sites; D002151:Calorimetry; D007704:Kininogens / Q000737:chemistry* / Q000378:metabolism; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism; D011288:Prekallikrein / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence; D013816:Thermodynamics; D014364:Tryptophan
D007704:Kininogens; D010446:Peptide Fragments; D014364:Tryptophan; D011288:Prekallikrein