Evaluation of phage display system and leech-derived tryptase inhibitor as a tool for understanding the serine proteinase specificities

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• 作者：Campos ; Ivan T.N. ; Silva ; Melissa M. ; Azzolini ; Simone S. ; Souza ; Adriana F. ; Sampaio ; Claudio A.M. ; et. al.
• 关键词：Phage display system ; Neutrophil elastase inhibitor ; Plasmin inhibitor ; Combinatorial library ; Kazal-type serine proteinase inhibitor ; Filamentous phage
• 刊名：Archives of Biochemistry and Biophysics
• 出版年：2004
• 期刊代码：116_00039861
• 类别：ch
• 出版时间：May 1, 2004
• 卷：425
• 期：1
• 页码：87-94
• 文件大小：134 K

摘要

A small combinatorial library of LDTI mutants (5.2×10⁴) restricted to the P1–P4^′ positions of the reactive site was displayed on the pCANTAB 5E phagemid, and LDTI fusion phages were produced and selected for potent neutrophil elastase and plasmin inhibitors. Strong fusion phage binders were analyzed by ELISA on enzyme-coated microtiter plates and the positive phages had their DNA sequenced. The LDTI variants: 29E (K8A, I9A, L10F, and K11F) and 19E (K8A, K11Q, and P12Y) for elastase and 2Pl (K11W and P12N), 8Pl (I9V, K11W, and P12E), and 10Pl (I9T, K11L, and P12L) for plasmin were produced with a Saccharomyces cerevisiae expression system. New strong elastase and plasmin inhibitors were 29E and 2Pl, respectively. LDTI-29E was a potent and specific neutrophil elastase inhibitor (K_i