Combined computational and structural studies to create novel macromolecular recognition properties

结合计算和结构研究来创造新的大分子识别特性

• 批准号: 10543489
• 负责人: BARRY L. STODDARD
• 金额: $ 35.2万
• 依托单位: FRED HUTCHINSON CANCER CENTER
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10543489
• 关键词: Affinity; Algorithms; Amino Acid Sequence; Base Pairing; Behavior; Binding; Binding Proteins; Binding Sites; Biochemical; Biological; Blinded; Collaborations; Complex; Complex Mixtures; Computer Analysis; Computer Models; Crystallography; DNA; DNA Binding; DNA-Binding Proteins; Dissection; Docking; Drug Design; Electrostatics; Engineering; Equilibrium; Hybrids; Hydrogen Bonding; Ligand Binding; Ligands; Modeling; Molecular; Motivation; Mutagenesis; Performance; Process; Property; Protein Conformation; Protein Engineering; Proteins; Protocols documentation; Resolution; Reverse engineering; Running; Sampling; Scaffolding Protein; Series; Solvents; Specificity; Structure; Surface; System; Tandem Repeat Sequences; Testing; Variant; Work; X-Ray Crystallography; data resource; design; design verification; improved; novel; pressure; protein complex; protein folding; protein structure; small molecule; statistical and machine learning

PROJECT SUMMARY The design of macromolecular binding interactions and complexes, and corresponding alteration of binding specificity, is a challenging endeavor that remains recalcitrant to computational approaches. This is true both for the creation of protein-protein complexes (which are driven by a enthalpic changes established primarily by stereochemical complementarity, balanced against large competing entropic changes) and for the redesign of protein-DNA complexes (which are heavily dependent upon DNA bending, hydrogen-bonds, electrostatic contacts, and the presence of solvent and counterions throughout the molecular interface). Over the past several years we have collaborated with several computational groups to help develop and validate computational approaches for the design and optimization of protein-protein recognition, protein-DNA recognition, and protein-small molecule recognition. Those studies have contributed to several new computational engineering approaches, including hybrid strategies that combine ab initio design of protein folds and binding sites, the ‘Rotamer Interaction Feld’ (RIF) docking protocol for efficient sampling of protein sequence and conformation, and novel parametric design approaches to create new tandem repeat proteins. We propose to continue this work through two specific aims to further develop and improve upon computational approaches for protein design. As part of this project, we will solve atomic resolution crystal structures of many selected and designed molecular complexes and provide them to our immediate collaborators as well as to a public structure prediction project, for computational prediction challenges. Aim 1. We will design and characterize novel self-associating circular tandem repeat proteins (using both de novo computational design and using high-throughput selections) and then further design them to undergo ligand-induced protein-protein association. Beyond the challenge of combining protein scaffold design and ligand binding design, the motivation for this aim is to determine the structural and mechanistic features of small molecule ligand-binding, and balance of forces, that facilitate ligand-induced protein-protein association. Aim 2. We will improve our understanding and ability to design novel protein-DNA recognition specificities and behaviors. To accomplish this, we will: (1) Systematically select and optimize a series of variants of a model DNA-binding protein, that display altered binding specificity across two regions of partially overlapping sequential clusters of basepairs and neighboring protein residues. (2) Determine the high-resolution structures and binding behavior of each construct. (3) Supervise blinded computational efforts, using multiple approaches, to predict the same structures. (4) Compare and analyze the results of computational predictions versus multiple computational prediction strategies to define features influencing predictive accuracy. For both aims, we will further exploit our crystallographic structures by computationally ‘reverse engineering’ each construct using validated protein structures, to further understand the performance of design approaches.

项目摘要 大分子结合相互作用和复合物的设计，以及结合的相应改变 特异性是一项挑战，仍然是计算方法的顽固性。这是真的 为了创建蛋白质 - 蛋白质复合物（由主要由焓变驱动 立体化学互补性，与大型竞争熵变化保持平衡）和重新设计 蛋白质-DNA复合物（在很大程度上取决于DNA弯曲，氢键，静电 触点，以及整个分子界面的溶剂和柜台的存在）。 在过去的几年中，我们与几个计算小组合作，以帮助发展和 验证蛋白质 - 蛋白质识别蛋白-DNA的设计和优化的计算方法 识别和蛋白质 - 小分子识别。这些研究促成了几个新的 计算工程方法，包括结合蛋白质概念设计的混合策略 折叠和结合位点，“ Rotamer相互作用FELD”（RIF）对接方案，用于有效抽样蛋白质 序列和构象以及新的参数设计方法，以创建新的串联重复蛋白。 我们建议通过两个特定的目标继续这项工作，以进一步发展和改进 蛋白质设计的计算方法。作为该项目的一部分，我们将解决原子分辨率晶体 许多选择和设计的分子络合物的结构，并将其提供给我们的直接 合作者以及公共结构预测项目以及计算预测挑战。 目的1。我们将设计和表征新型的自缔约圆形串联重复蛋白（同时使用DE Novo计算设计并使用高通量选择），然后进一步设计以进行 配体诱导的蛋白质 - 蛋白质关联。除了结合蛋白质脚手架设计和 配体结合设计，此目的的动机是确定 小分子配体结合和力平衡，促进了配体诱导的蛋白质蛋白缔合。 目标2。我们将提高我们设计新型蛋白质DNA识别规范的理解和能力 行为。为此，我们将：（1）系统选择并优化模型的一系列变体 DNA结合蛋白，在部分重叠的两个区域中显示出改变的结合特异性 底培和相邻蛋白质的顺序簇保留。 （2）确定高分辨率结构 和每个结构的绑定行为。 （3）使用多个监督盲目的计算工作 方法，以预测相同的结构。 （4）比较和分析计算预测的结果 与多个计算预测策略的定义特征会影响预测精度。 对于这两个目标，我们将通过计算“反向工程”进一步利用我们的晶体结构 使用经过验证的蛋白质结构的每个结构，以进一步了解设计方法的性能。

项目成果

De novo design of protein homodimers containing tunable symmetric protein pockets.

• DOI: 10.1073/pnas.2113400119
• 发表时间: 2022-07-26
• 期刊: Proceedings of the National Academy of Sciences of the United States of America
• 影响因子: 11.1

Design of functionalised circular tandem repeat proteins with longer repeat topologies and enhanced subunit contact surfaces.

• DOI: 10.1038/s42003-021-02766-y
• 发表时间: 2021-10-29
• 期刊: Communications biology
• 影响因子: 5.9
• 作者: Hallinan JP;Doyle LA;Shen BW;Gewe MM;Takushi B;Kennedy MA;Friend D;Roberts JM;Bradley P;Stoddard BL
• 通讯作者: Stoddard BL

Stepwise design of pseudosymmetric protein hetero-oligomers.

假对称蛋白质异源寡聚物的逐步设计。

• DOI: 10.1101/2023.04.07.535760
• 发表时间: 2023
• 期刊: bioRxiv : the preprint server for biology
• 作者: Kibler,RyanD;Lee,Sangmin;Kennedy,MadisonA;Wicky,BasileIM;Lai,StellaM;Kostelic,MariusM;Li,Xinting;Chow,CameronM;Carter,Lauren;Wysocki,VickiH;Stoddard,BarryL;Baker,David
• 通讯作者: Baker,David

De novo design of knotted tandem repeat proteins.

• DOI: 10.1038/s41467-023-42388-y
• 发表时间: 2023-10-24
• 期刊: NATURE COMMUNICATIONS
• 影响因子: 16.6
• 作者: Doyle, Lindsey A.;Takushi, Brittany;Kibler, Ryan D.;Milles, Lukas F.;Orozco, Carolina T.;Jones, Jonathan D.;Jackson, Sophie E.;Stoddard, Barry L.;Bradley, Philip
• 通讯作者: Bradley, Philip

Immunization with a self-assembling nanoparticle vaccine displaying EBV gH/gL protects humanized mice against lethal viral challenge.

• DOI: 10.1016/j.xcrm.2022.100658
• 发表时间: 2022-06-21
• 期刊: CELL REPORTS MEDICINE
• 影响因子: 14.3
• 作者: Malhi, Harman;Homad, Leah J.;Wan, Yu-Hsin;Poudel, Bibhav;Fiala, Brooke;Borst, Andrew J.;Wang, Jing Yang;Walkey, Carl;Price, Jason;Wall, Abigail;Singh, Suruchi;Moodie, Zoe;Carter, Lauren;Handa, Simran;Correnti, Colin E.;Stoddard, Barry L.;Veesler, David;Pancera, Marie;Olson, James;King, Neil P.;McGuire, Andrew T.
• 通讯作者: McGuire, Andrew T.