Molecular Brain Research Group, Robarts Research Institute, Department of Anatomy & Cell Biology, Schulich School of Medicine & Dentistry, University of Western Ontario, Canada.
Toxicon. 2012 Oct;60(5):907-18. doi: 10.1016/j.toxicon.2012.05.026. Epub 2012 May 31.
PnTx3-4 is a toxin isolated from the venom of the spider Phoneutria nigriventer that blocks N-, P/Q-, and R-type voltage-gated calcium channels and has great potential for clinical applications. In this report we used the SUMO system to express large amounts of recombinant PnTx3-4 peptide, which was found in both soluble and insoluble fractions of bacterial extracts. We purified the recombinant toxin from both fractions and showed that the recombinant peptide showed biological activity similar to the native PnTx3-4. In silico analysis of the primary sequence of PnTx3-4 indicated that the peptide conforms to all the criteria of a knottin scaffold. Additionally, circular dichroism spectrum analysis of the recombinant PnTx3-4 predicted that the toxin structure is composed of approximately 53% turns/unordered, 31% α-helix and 16% β-strand, which is consistent with predicted model of the PnTx3-4 knottin scaffold available at the knottin database (http://knottin.cbs.cnrs.fr). These studies provide the basis for future large scale production and structure-function investigation of PnTx3-4.
PnTx3-4 是从蜘蛛 Phoneutria nigriventer 的毒液中分离出来的一种毒素，它可以阻断 N-、P/Q- 和 R-型电压门控钙通道，具有很大的临床应用潜力。在本报告中，我们使用 SUMO 系统表达了大量的重组 PnTx3-4 肽，该肽存在于细菌提取物的可溶性和不溶性部分中。我们从这两个部分纯化了重组毒素，并表明重组肽表现出与天然 PnTx3-4 相似的生物活性。PnTx3-4 的一级序列的计算机分析表明，该肽符合所有 knottin 支架的标准。此外，对重组 PnTx3-4 的圆二色性光谱分析预测，该毒素结构由大约 53%的环/无序、31%的α-螺旋和 16%的β-折叠组成，这与 knottin 数据库中可用的 PnTx3-4 knottin 支架的预测模型一致(http://knottin.cbs.cnrs.fr)。这些研究为未来 PnTx3-4 的大规模生产和结构功能研究提供了基础。
