Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario, Canada.
PLoS Comput Biol. 2010 Dec 9;6(12):e1001029. doi: 10.1371/journal.pcbi.1001029.
Chelt, a cholera-like toxin from Vibrio cholerae, and Certhrax, an anthrax-like toxin from Bacillus cereus, are among six new bacterial protein toxins we identified and characterized using in silico and cell-based techniques. We also uncovered medically relevant toxins from Mycobacterium avium and Enterococcus faecalis. We found agriculturally relevant toxins in Photorhabdus luminescens and Vibrio splendidus. These toxins belong to the ADP-ribosyltransferase family that has conserved structure despite low sequence identity. Therefore, our search for new toxins combined fold recognition with rules for filtering sequences--including a primary sequence pattern--to reduce reliance on sequence identity and identify toxins using structure. We used computers to build models and analyzed each new toxin to understand features including: structure, secretion, cell entry, activation, NAD+ substrate binding, intracellular target binding and the reaction mechanism. We confirmed activity using a yeast growth test. In this era where an expanding protein structure library complements abundant protein sequence data--and we need high-throughput validation--our approach provides insight into the newest toxin ADP-ribosyltransferases.
霍乱弧菌中的霍乱毒素样蛋白 Chelt 和蜡状芽孢杆菌中的炭疽毒素样蛋白 Certhrax 是我们使用计算机模拟和基于细胞的技术鉴定和表征的六种新型细菌蛋白毒素。我们还从鸟分枝杆菌和粪肠球菌中发现了具有医学相关性的毒素。我们在发光光杆藻和灿烂弧菌中发现了具有农业相关性的毒素。这些毒素属于 ADP-核糖基转移酶家族，尽管序列相似性较低，但具有保守的结构。因此，我们的新毒素搜索将折叠识别与用于过滤序列的规则（包括原始序列模式）相结合，以减少对序列相似性的依赖，并使用结构来识别毒素。我们使用计算机构建模型并分析每个新毒素，以了解包括结构、分泌、细胞进入、激活、NAD+ 底物结合、细胞内靶标结合和反应机制在内的特征。我们使用酵母生长测试来验证其活性。在这个蛋白质结构库不断扩展、蛋白质序列数据丰富的时代——我们需要高通量验证——我们的方法为最新的毒素 ADP-核糖基转移酶提供了深入了解。
