Markus G, DePasquale J L, Wissler F C
J Biol Chem. 1978 Feb 10;253(3):727-32.
The binding of epsilon-amino[14C]caproic acid (6-aminohexanoic acid, EACA) to native human plasminogen was determined using the ultrafiltration technique of Paulus (Paulus, H. (1969) Anal. Biochem. 32, 91-100) at free ligand concentrations ranging from 2 micrometer to 16 mM. One strong binding site (Kd = 0.009 mM) and approximately five weaker ones (Kd = 5 mM) were found. The constants were obtained by fitting the experimental points to the simple assumption of two sets of noninteracting sites. The distinct separation of the two kinds of sites allowed the correlation of the well known epsilon-aminocaproic acid-induced conformational transition in plasminogen with the saturation of the weaker group of binding sites by this ligand. The conformational transition was monitored by measurements of the sedimentation coefficient, as was done by others earlier. The midpoint of the transition occurred at approximately 3.3 mM free ligand. A dissociation constant of 0.32 mM was also obtained for L-lysine by measurements of the competition between this compound and labeled epsilon-aminocaproic acid for the strong binding site. The correlation between epsilon-aminocaproic acid binding and effects of the compound on various physical and functional properties is discussed. A discussion of the possible sources of error encountered in the technique used is also included.
采用保卢斯的超滤技术（Paulus, H. (1969) Anal. Biochem. 32, 91 - 100），在游离配体浓度范围为2微摩尔至16毫摩尔的条件下，测定了ε-氨基[¹⁴C]己酸（6-氨基己酸，EACA）与天然人纤溶酶原的结合情况。发现了一个强结合位点（Kd = 0.009毫摩尔）和大约五个较弱的结合位点（Kd = 5毫摩尔）。这些常数是通过将实验点拟合到两组非相互作用位点的简单假设而获得的。两种位点的明显分离使得纤溶酶原中众所周知的ε-氨基己酸诱导的构象转变与该配体对较弱结合位点组的饱和作用相关联。如前人所做的那样，通过测量沉降系数来监测构象转变。转变的中点出现在约3.3毫摩尔游离配体处。通过测量该化合物与标记的ε-氨基己酸对强结合位点的竞争作用，还获得了L-赖氨酸的解离常数为0.32毫摩尔。讨论了ε-氨基己酸结合与该化合物对各种物理和功能特性的影响之间的相关性。还包括了对所用技术中可能遇到的误差来源的讨论。
