CAVER 3.0: A Tool for the Analysis of Transport Pathways in Dynamic Protein Structures

Tunnels and channels facilitate the transport of small molecules, ions and water solvent in a large variety of proteins. Characteristics of individual transport pathways, including their geometry, physico-chemical properties and dynamics are instrumental for understanding of structure-function relationships of these proteins, for the design of new inhibitors and construction of improved biocatalysts. CAVER is a software tool widely used for the identification and characterization of transport pathways in static macromolecular structures. Herein we present a new version of CAVER enabling automatic analysis of tunnels and channels in large ensembles of protein conformations. CAVER 3.0 implements new algorithms for the calculation and clustering of pathways. A trajectory from a molecular dynamics simulation serves as the typical input, while detailed characteristics and summary statistics of the time evolution of individual pathways are provided in the outputs. To illustrate the capabilities of CAVER 3.0, the tool was applied for the analysis of molecular dynamics simulation of the microbial enzyme haloalkane dehalogenase DhaA. CAVER 3.0 safely identified and reliably estimated the importance of all previously published DhaA tunnels, including the tunnels closed in DhaA crystal structures. Obtained results clearly demonstrate that analysis of molecular dynamics simulation is essential for the estimation of pathway characteristics and elucidation of the structural basis of the tunnel gating. CAVER 3.0 paves the way for the study of important biochemical phenomena in the area of molecular transport, molecular recognition and enzymatic catalysis. The software is freely available as a multiplatform command-line application at http://www.caver.cz.各类蛋白质内部的隧道与通道可介导小分子、离子及水性溶剂的转运过程。各类转运通路的自身特征，包括其几何形态、理化性质与动态变化，对于解析此类蛋白质的结构-功能关系、设计新型抑制剂以及构建优化的生物催化剂均具有关键作用。CAVER是一款被广泛应用于静态大分子结构中转运通路识别与特征表征的软件工具。本文介绍了全新版本的CAVER，其可实现对大规模蛋白质构象集合中隧道与通道的自动化分析。CAVER 3.0新增了用于通路计算与聚类的全新算法。该软件的典型输入为分子动力学（molecular dynamics）模拟轨迹，输出结果则包含各通路随时间演化的详细特征与汇总统计数据。为验证CAVER 3.0的性能，本研究将其应用于微生物酶卤代烷脱卤酶DhaA（haloalkane dehalogenase DhaA）的分子动力学模拟分析。CAVER 3.0成功识别出此前文献中报道的所有DhaA相关隧道，并可靠评估了各隧道的重要性，其中包括DhaA晶体结构中处于闭合状态的隧道。所得结果清晰表明，分子动力学模拟分析对于准确估算通路特征、阐明隧道门控的结构基础不可或缺。CAVER 3.0为分子转运、分子识别及酶催化领域内重要生化现象的研究铺平了道路。该软件作为一款支持多平台的命令行应用程序，可通过http://www.caver.cz免费获取。