Recombinant production, purification and biochemical characterization of domain 6 of LEKTI: a temporary Kazal-type-related serine proteinase inhibitor

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• 作者：Kreutzmann ; Peter ; Schulz ; Axel ; Stä ; ndker ; Ludger ; Forssmann ; Wolf-Georg ; Mä ; gert ; Hans-Jü ; rgen
• 关键词：LD-6 ; LEKTI domain 6 ; EK ; enterokinase ; HF7665 ; hemofiltrate peptide with a molecular weight of 7665Da ; LEKTI ; lympho-epithelial Kazal-type-related inhibitor ; CZE ; capillary zone electrophoresis ; l-BAPNA ; Nα ; -benzoyl-l-arginine p-nitroanilide ; Recombi
• 刊名：Journal of Chromatography B, Analytical Technologies in the Biomedical and Life Sciences
• 出版年：2004
• 出版时间：April 15, 2004
• 年：2004
• 卷：803
• 期：1
• 页码：75-81
• 全文大小：160 K

文摘

Lympho-epithelial Kazal-type-related inhibitor (LEKTI) is a 15-domain serine proteinase inhibitor which is of pathophysiological relevance for skin diseases and atopy. Domains 2 and 15 of LEKTI contain six cysteine residues and match the Kazal-type inhibitor motif almost exactly. The other 13 domains seem to be Kazal-type derived but lack the cysteines in positions 3 and 6 usually conserved within this family of inhibitors. Here, we report the recombinant production and comprehensive biochemical characterization of the 7.7kDa LEKTI domain 6 (LD-6). Testing a selected number of different serine proteinases, we show that both native and recombinant LD-6 exhibit a significant but temporary inhibitory activity on trypsin. Furthermore, the relation of LEKTI domain 6 to Kazal-type inhibitors is confirmed by determining its disulfide bond pattern (1–4/2–3) and its P₁ site located after the second Cys residue of LD-6. The established strategy for the recombinant production of LEKTI domain 6 will enable further investigation of its mode of action and its physiological role.