Binnie C G, Lord S T
Department of Pathology, University of North Carolina, Chapel Hill 27599.
Thromb Haemost. 1991 Feb 12;65(2):165-8.
Binding of the synthetic peptide AAKDSDWPFAS-DEDWNYKAPSGAR, a fibrinogen alpha 27-50 analog, to thrombin was studied by inhibition assays and affinity chromatography. Peptide alpha 27-50 corresponds to a segment of human fibrinogen downstream from the thrombin cleavage site, with cysteine residues at positions 28, 36, 45 and 49 replaced by alanine. The peptide inhibited clotting of fibrinogen with an inhibition constant of 190-400 microM. Cleavage of fibrinopeptides A and B was inhibited by the peptide and the peptide was competitive with fibrinogen for thrombin. Inhibition of the small substrate tosyl-Gly-Pro-Arg-p-nitroaniline was not observed indicating that the peptide did not block the active site of the enzyme. Peptide alpha 27-50 that was covalently linked to Sepharose bound active site-inhibited thrombin at low ionic strength and was eluted at higher salt concentration. The peptide was not cleaved on overnight exposure to thrombin as determined by reverse phase HPLC. In summary, the peptide bound to, but was not a substrate for thrombin. These results suggest that this region of fibrinogen contributes to binding of thrombin.
通过抑制试验和亲和色谱法研究了合成肽AAKDSDWPFAS - DEDWNYKAPSGAR（一种纤维蛋白原α27 - 50类似物）与凝血酶的结合。肽α27 - 50对应于人纤维蛋白原凝血酶切割位点下游的一段序列，其中28、36、45和49位的半胱氨酸残基被丙氨酸取代。该肽抑制纤维蛋白原的凝血，抑制常数为190 - 400微摩尔。该肽抑制纤维蛋白肽A和B的裂解，并且该肽与纤维蛋白原竞争凝血酶。未观察到对小底物甲苯磺酰 - 甘氨酰 - 脯氨酰 - 精氨酰 - 对硝基苯胺的抑制作用，表明该肽未阻断酶的活性位点。与琼脂糖共价连接的肽α27 - 50在低离子强度下结合活性位点被抑制的凝血酶，并在较高盐浓度下洗脱。通过反相高效液相色谱法测定，该肽在与凝血酶过夜孵育后未被裂解。总之，该肽与凝血酶结合，但不是凝血酶的底物。这些结果表明纤维蛋白原的这一区域有助于凝血酶的结合。
