Filippusson H, Erlendsson L S, Lowe C R
Department of Biochemistry, Science Institute, University of Iceland, Reykjavík.
J Mol Recognit. 2000 Nov-Dec;13(6):370-81. doi: 10.1002/1099-1352(200011/12)13:6<370::AID-JMR510>3.0.CO;2-5.
A low-molecular-weight biomimetic affinity ligand selective for binding elastase has been designed and synthesized. The ligand was based on mimicking part of the interaction between a natural inhibitor, turkey ovomucoid inhibitor and elastase, and modelled from the X-ray crystallographic structure of the enzyme-inhibitor complex. Limited solid-phase combinatorial chemistry was used to synthesize 12 variants of the lead ligand using the triazine moiety as the scaffold for assembly. The ligand library was screened for its ability to bind elastase and trypsin, and two ligands were studied further. Ligand C4/6 [2-alanyl-alanyl-4-tryptamino-6-(alpha-lysyl)-s-triazine] was found to bind porcine pancreatic elastase, but not trypsin, with a dissociation constant of 6 x 10(-5) M and a binding capacity of 21 mg elastase per ml gel. The adsorbent was used to purify elastase from a crude extract of porcine pancreas. Immobilized ligand C4/5 6 [2-alanyl-alanyl-4-tyramino-6-(alpha-lysyl)-s-triazine] was similarly chosen for optimal binding of elastase from cod and used to purify the enzyme from a crude extract of cod pyloric caeca. Ligand C4/6 was subsequently synthesized in solution and its structure verified by 1H-NMR.
一种对弹性蛋白酶具有选择性结合作用的低分子量仿生亲和配体已被设计并合成出来。该配体基于模拟天然抑制剂——火鸡卵类粘蛋白抑制剂与弹性蛋白酶之间的部分相互作用，并根据酶 - 抑制剂复合物的X射线晶体结构进行建模。利用有限的固相组合化学方法，以三嗪部分作为组装支架，合成了12种先导配体变体。对配体库结合弹性蛋白酶和胰蛋白酶的能力进行了筛选，并对两种配体进行了进一步研究。发现配体C4/6 [2 - 丙氨酰 - 丙氨酰 - 4 - 色氨酰 - 6 -（α - 赖氨酰）- s - 三嗪]能结合猪胰弹性蛋白酶，但不结合胰蛋白酶，其解离常数为6×10⁻⁵ M，结合容量为每毫升凝胶结合21毫克弹性蛋白酶。该吸附剂用于从猪胰腺粗提物中纯化弹性蛋白酶。固定化配体C4/5 6 [2 - 丙氨酰 - 丙氨酰 - 4 - 酪氨酰 - 6 -（α - 赖氨酰）- s - 三嗪]同样被选择用于从鳕鱼中最佳结合弹性蛋白酶，并用于从鳕鱼幽门盲囊粗提物中纯化该酶。随后在溶液中合成了配体C4/6，并通过¹H - NMR验证了其结构。
