Boosani Chandra Shekhar, Sudhakar Akulapalli
Cell Signaling and Angiogenesis Laboratory, Department of Genetics, Boys Town National Research Hospital, Omaha, NE, USA.
Protein Expr Purif. 2006 Oct;49(2):211-8. doi: 10.1016/j.pep.2006.03.007. Epub 2006 Apr 3.
alpha1(IV)NC1, a cleavage fragment of the carboxy terminal non-collagenous human alpha1 chain of type IV collagen, is derived from the extracellular matrix specifically by MMP-2. Recently we determined the in vitro and in vivo anti-angiogenic activity of alpha1(IV)NC1 and presently, its role in cancer therapy is under evaluation. To characterize alpha1(IV)NC1 as a potential candidate for drug development and to test its efficacy in animal models, an effective method to produce a purified active form of alpha1(IV)NC1 is needed. In the present study, expression of alpha1(IV)NC1 in Sf9 cells using baculovirus expression system was discussed, this method was found to be effective in the production of a functionally active soluble form of the recombinant protein. The purified protein showed its characteristic activities such as inhibiting cell proliferation, migration, and tube formation in endothelial cells.
α1（IV）NC1是IV型胶原蛋白的人α1链羧基末端非胶原蛋白的裂解片段，它特别由基质金属蛋白酶-2从细胞外基质中产生。最近我们确定了α1（IV）NC1的体外和体内抗血管生成活性，目前，其在癌症治疗中的作用正在评估中。为了将α1（IV）NC1表征为药物开发的潜在候选物并在动物模型中测试其功效，需要一种有效的方法来生产纯化的活性形式的α1（IV）NC1。在本研究中，讨论了使用杆状病毒表达系统在Sf9细胞中表达α1（IV）NC1，发现该方法在生产重组蛋白的功能活性可溶性形式方面是有效的。纯化的蛋白显示出其特征活性，如抑制内皮细胞的增殖、迁移和管形成。
