Semenova O A, Strukova S M
Biokhimiia. 1980 Dec;45(12):2225-32.
The interaction of alpha- and beta gamma-thrombin with heparin was studied by ultraviolet difference spectroscopy within the wavelength range of 230-300 nm. The absorption difference spectrum of the thrombin-heparin complex was negative and had two maxima at 255 nm (5300 M-1 cm-1) and 282 nm (4700 M-1 cm-1) for alpha-thrombin and at 240 nm (4900 M-1 cm-1) and 282 nm (4100 M-1 cm-1) for beta gamma-thrombin. It is assumed that the conformational changes induced by heparin in the enzyme molecule involve the transfer of some tryptophan and tyrosine residues from the interior of the protein to the surface. The absorption changes during alpha-thrombin--heparin interaction at physiological ionic strength suggest binding of some alpha-thrombin molecules to a heparin molecule at the ligand-enzyme molar ratio lower than 1. Under the same conditions beta gamma-thrombin forms an equimolar complex with heparin with the dissociation constant equal to 7,0.10(-9) M. The ionic strength increase up to 0,217 M NaCl results in some disturbances in beta gamma-thrombin-heparin interaction and prevents the binding of additional alpha-thrombin molecules to an equimolar complex of alpha-thrombin with heparin. Therefore the kinetics of the two enzyme forms interaction with heparin are similar, the alpha-thrombin affinity for heparin being a little higher. The data obtained suggest that alpha-thrombin binding to heparin is essential for biological inactivation of thrombin.
在230 - 300nm波长范围内，采用紫外差示光谱法研究了α-凝血酶和βγ-凝血酶与肝素的相互作用。凝血酶-肝素复合物的吸收差光谱为负，α-凝血酶在255nm（5300 M-1 cm-1）和282nm（4700 M-1 cm-1）处有两个最大值，βγ-凝血酶在240nm（4900 M-1 cm-1）和282nm（4100 M-1 cm-1）处有两个最大值。据推测，肝素在酶分子中诱导的构象变化涉及一些色氨酸和酪氨酸残基从蛋白质内部转移到表面。在生理离子强度下，α-凝血酶与肝素相互作用期间的吸收变化表明，一些α-凝血酶分子在配体-酶摩尔比低于1时与肝素分子结合。在相同条件下，βγ-凝血酶与肝素形成等摩尔复合物，解离常数等于7×10^(-9) M。离子强度增加到0.217M NaCl会导致βγ-凝血酶与肝素的相互作用出现一些紊乱，并阻止额外的α-凝血酶分子与α-凝血酶与肝素的等摩尔复合物结合。因此，两种酶形式与肝素相互作用的动力学相似，α-凝血酶对肝素的亲和力略高。所得数据表明，α-凝血酶与肝素的结合对于凝血酶的生物学失活至关重要。
