Department of Clinical Chemistry, P.O. Box 63, University of Helsinki and Helsinki University Central Hospital, FIN-00014 University of Helsinki, Helsinki, Finland.
Department of Biosciences and Biocenter Kuopio, University of Kuopio, P.O. Box 1627, FIN-70211 Kuopio, Finland.
Biol Chem. 2010 Feb-Mar;391(2-3):283-293. doi: 10.1515/bc.2010.030.
Human trypsin isoenzymes share extensive sequence similarity, but certain differences in their activity and susceptibility to inhibitors have been observed. Using phage display technology, we identified seven different peptides that bind to and inhibit the activity of trypsin-3, a minor trypsin isoform expressed in pancreas and brain. All of the peptides contain at least two of the amino acids tryptophan, alanine and arginine, whereas proline was found closer to the N-terminus in all but one peptide. All peptides contain two or more cysteines, suggesting a cyclic structure. However, we were able to make synthetic linear variants of these peptides without losing bioactivity. Alanine replacement experiments for one of the peptides suggest that the IPXXWFR motif is important for activity. By molecular modeling the same amino acids were found to interact with trypsin-3. The peptides also inhibit trypsin-1, but only weakly, if at all, trypsin-2 and -C. As trypsin is a highly active enzyme which can activate protease-activated receptors and enzymes that participate in proteolytic cascades involved in tumor invasion and metastasis, these peptides might be useful lead molecules for the development of drugs for diseases associated with increased trypsin activity.
人胰蛋白酶同工酶具有广泛的序列相似性，但在活性和对抑制剂的敏感性方面存在某些差异。我们使用噬菌体展示技术鉴定了七个不同的肽段，这些肽段可以与胰蛋白酶-3结合并抑制其活性，胰蛋白酶-3是一种在胰腺和大脑中表达的次要胰蛋白酶同工酶。所有肽段至少含有两个氨基酸色氨酸、丙氨酸和精氨酸，而脯氨酸除一个肽段外，其余都更靠近 N 端。所有肽段都含有两个或更多的半胱氨酸，提示其具有环状结构。然而，我们能够合成这些肽段的线性变体而不丧失生物活性。对其中一个肽段的丙氨酸替换实验表明，IPXXWFR 基序对活性很重要。通过分子建模发现相同的氨基酸与胰蛋白酶-3相互作用。这些肽段还可以抑制胰蛋白酶-1，但如果有的话，抑制作用也很弱，对胰蛋白酶-2 和 -C 几乎没有抑制作用。由于胰蛋白酶是一种高度活跃的酶，可以激活蛋白酶激活受体和参与肿瘤侵袭和转移的蛋白水解级联反应的酶，因此这些肽段可能是开发与胰蛋白酶活性增加相关疾病药物的有用先导分子。
