Ito N, Noguchi K, Kazama M, Shimura K, Kasai K
J Chromatogr. 1985 Nov 27;348(1):199-204. doi: 10.1016/s0021-9673(01)92453-7.
Human Glu-plasminogen, Lys-plasminogen and plasmin were effectively separated by high-performance affinity chromatography. The affinity adsorbent was prepared by using a micro-particulate polyvinyl alcohol gel (Asahipak GS-gel) as the supporting material and p-aminobenzamidine as the specific ligand. All of the active enzyme and proenzymes were adsorbed. Glu-plasminogen was eluted by changing the pH of the eluent and Lys-plasminogen by using an eluent containing 6-amino-hexanoic acid. This affinity adsorbent recognized the difference between these proenzyme species. For the elution of plasmin, addition of urea was necessary. Plasmin may have been adsorbed through a two-site interaction with the adsorbent. All proteins were eluted as sharp peaks and the time required for one cycle was about 1 h. Fluorimetric detection of eluted protein and on-line assay of enzyme activity using a fluorigenic substrate made it possible to analyse microgram amounts of proteins specifically.
人谷氨酸纤溶酶原、赖氨酸纤溶酶原和纤溶酶通过高效亲和色谱法得到有效分离。亲和吸附剂是用微粒聚乙烯醇凝胶（旭pak GS凝胶）作为支撑材料，对氨基苯甲脒作为特异性配体制备而成。所有活性酶和酶原均被吸附。通过改变洗脱液的pH值洗脱谷氨酸纤溶酶原，使用含6-氨基己酸的洗脱液洗脱赖氨酸纤溶酶原。这种亲和吸附剂能够识别这些酶原种类之间的差异。为了洗脱纤溶酶，需要添加尿素。纤溶酶可能是通过与吸附剂的双位点相互作用而被吸附的。所有蛋白质均以尖锐峰的形式洗脱，一个循环所需时间约为1小时。通过荧光检测洗脱的蛋白质以及使用荧光底物对酶活性进行在线测定，可以特异性地分析微克量的蛋白质。
