De novo design of small-molecule-binding proteins
小分子结合蛋白的从头设计
基本信息
- 批准号:10217208
- 负责人:
- 金额:$ 9.94万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2020
- 资助国家:美国
- 起止时间:2020-07-15 至 2022-04-29
- 项目状态:已结题
- 来源:
- 关键词:AchievementAffinityAgreementAlgorithm DesignAmino Acid SequenceAmino AcidsAmino Acyl-tRNA SynthetasesAnticoagulantsAntidotesAntithrombin IIIAwardBindingBinding ProteinsBinding SitesChargeClinicalCoagulation ProcessCollaborationsComplementComplexComputer ModelsComputing MethodologiesCrystallizationDatabasesDevelopmentDrug Delivery SystemsEnzymesEventEvolutionGeometryGoalsHealthHeterogeneityHumanHydrogen BondingHydrophobicityIndividualKnowledgeLaboratoriesLearningLibrariesLifeLigandsLigaseMammalian CellMediatingMedicalMethodsModernizationMolecular BiologyMolecular ConformationMolecular ProbesMotivationMutagenesisNaturePharmaceutical PreparationsPharmacologic SubstancePhosphoserinePhysiciansPlayPorphyrinsPost-Translational Protein ProcessingPrincipal InvestigatorProcessProtein EngineeringProtein Structure DatabasesProteinsPublicationsRecording of previous eventsResearchResearch TrainingResolutionRoentgen RaysRoleSamplingScheduleScientistSet proteinSpecificityStructureSystemTemperatureTestingTimeTrainingVariantVertebral columnWaterWorkX-Ray Crystallographybaseclinical applicationclinically relevantcombinatorialdata repositorydesignfunctional groupinfancyinsightmeetingsmodel designmolecular dynamicsmolecular recognitionmutantnovelphosphohistidineporphyrin aprofessorprogramsprotein data bankprotein foldingprotein functionprotein protein interactionpyrrolysinescreeningsensorsmall moleculesuccesstyrosine O-sulfateunnatural amino acidswater sampling
项目摘要
PROJECT SUMMARY
Protein-ligand binding events underlay all life processes. Protein design tests and extends our knowledge of
protein folding and function through the creation of proteins from scratch. This proposal aims to develop a
computational method for the design of proteins that bind to any small molecule with high affinity and
selectivity. The state-of-the-art in ligand-binding protein design critically relies on random experimental
optimization and screening. If we truly understand how proteins bind small molecules, we should be able to go
directly from computer models to tight binders. The hypothesis that drives this proposal is that proteins use a
vast but now enumerable number of molecular interaction motifs combinatorially throughout evolution to create
the binding sites of modern-day proteins. Computational methods will be employed to uncover this set of
interactions in the large database of protein structures available in the protein databank (PDB). Binding sites
will be designed by sampling motifs for all functional groups of a ligand onto a protein backbone. We call this
design method Convergent Motifs for Binding Sites (COMBS). COMBS was used to design ABLER, the first
ligand-binding protein designed from scratch to bind its target ligand—the antithrombotic drug apixaban—with
an unprecedentedly high affinity, without experimental optimization of sequence. ABLER has potential clinical
relevance as an anti-clotting antidote, although that is outside the scope of the proposal. High-resolution crystal
structures of ABLER agree with the design model, both in overall topology and the intended molecular
interactions with the ligand. Aim 1 of this proposal focuses on designing variants of ABLER to increase affinity
and probe the molecular bases for the observed drug-protein interactions. Aim 2 focuses on the role of water in
ligand-binding protein design, motivated by the water-mediated protein-ligand interaction found in the crystal
structure. In this Aim, I will curate a database of water-protein interactions from the PDB and use these to
sample water-mediated protein-ligand interactions during design. I will also learn to use explicit-water
molecular dynamics simulations to critically assess the roles of water in binding. Aim 3 uses COMBS to redesign
the binding site of pyrrolysine tRNA synthetase for incorporation of charged unnatural amino acids (such as
sulfotyrosine) into mammalian cells, since laboratory evolution and library screens for this goal have so far been
unsuccessful. These aims will augment my training in molecular biology, computational protein design, and
protein structural characterization (X-ray crystallography and NMR). The K99 portion of this work in the
DeGrado lab will expose me to all aspects of the scientific process, from inception to publication. Bill is a world
expert in protein design, and his insight is critical to the success of the project. At UCSF, I will gain much through
my regularly scheduled meetings with Ethan Weiss, who brings the perspective of a physician scientist with a
long history of antithrombotic research and clinical applications. My collaboration with UCSF professor Lei
Wang will expose me to the field of unnatural amino acid incorporation and will be critical for applying COMBS
to the most impactful targets for mimics of post-translational modifications. The research and training proposed
herein will greatly complement my current skillset and background, which will be essential to my research
program as I transition into an independent principal investigator.
项目总结
蛋白质-配体结合事件是所有生命过程的基础。蛋白质设计测试并扩展了我们对
蛋白质折叠和功能是通过从头开始创造蛋白质来实现的。这项提案旨在开发一种
设计与任何高亲和力和高亲和力的小分子结合的蛋白质的计算方法
选择性。最先进的配体结合蛋白设计在很大程度上依赖随机实验
优化筛选。如果我们真正了解蛋白质是如何结合小分子的,我们应该能够
直接从计算机模型到紧密的活页夹。推动这一提议的假设是蛋白质使用一种
在整个进化过程中,大量但现在可以计数的分子相互作用基序组合在一起,创造了
现代蛋白质的结合部位。将使用计算方法来揭示这一组
蛋白质数据库(PDB)中提供的大型蛋白质结构数据库中的相互作用。结合部位
将通过在蛋白质骨架上采样配体的所有官能团的基序来设计。我们把这叫做
设计方法结合位点的趋同基序(梳子)。Combs被用于设计Enabler,第一个
从头开始设计的配体结合蛋白,用于将其靶标配体-抗血栓药物阿皮沙班-与
一种前所未有的高亲和力,没有实验的序列优化。Enabler具有潜在的临床应用前景
相关性作为一种抗凝血解毒剂,尽管这不在提案的范围内。高分辨率晶体
Abler的结构与设计模型一致,无论是总体拓扑结构还是目标分子
与配基的相互作用。该提案的目标1侧重于设计Enabler的变体以增加亲和力
并探索观察到的药物-蛋白质相互作用的分子基础。目标2侧重于水在
在晶体中发现的水介导的蛋白质-配体相互作用的激励下的配体结合蛋白设计
结构。在这个目标中,我将从PDB建立一个水-蛋白质相互作用的数据库,并使用这些数据库来
设计过程中水介导的蛋白质-配基相互作用样本。我还将学习使用显性水
分子动力学模拟,以严格评估水在结合中的作用。Aim 3使用梳子重新设计
吡咯赖氨酸tRNA合成酶与带电非天然氨基酸(如
硫代酪氨酸)进入哺乳动物细胞,因为到目前为止,这个目标的实验室进化和文库筛选
不成功。这些目标将加强我在分子生物学、计算蛋白质设计和
蛋白质结构表征(X射线结晶学和核磁共振)。这项工作的K99部分在
降级实验室将使我接触到科学过程的方方面面,从开始到发表。比尔就是一个世界
他是蛋白质设计方面的专家,他的洞察力对项目的成功至关重要。在加州大学旧金山分校,我将通过
我定期与Ethan Weiss会面,他带来了一位内科科学家的观点,
具有悠久的抗血栓研究和临床应用历史。我与加州大学旧金山分校雷教授的合作
王将让我接触到非天然氨基酸结合的领域,并将是应用梳子的关键
对最有影响力的目标进行翻译后修改的模仿。建议的研究和培训
这将极大地补充我目前的技能和背景,这对我的研究是必不可少的
当我转变为一名独立的首席调查员时。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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Nicholas Polizzi其他文献
Nicholas Polizzi的其他文献
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{{ truncateString('Nicholas Polizzi', 18)}}的其他基金
De novo design of small-molecule-binding proteins
小分子结合蛋白的从头设计
- 批准号:
10683406 - 财政年份:2020
- 资助金额:
$ 9.94万 - 项目类别:
De novo design of small-molecule-binding proteins
小分子结合蛋白的从头设计
- 批准号:
10055537 - 财政年份:2020
- 资助金额:
$ 9.94万 - 项目类别:
De novo design of small-molecule-binding proteins
小分子结合蛋白的从头设计
- 批准号:
10604467 - 财政年份:2020
- 资助金额:
$ 9.94万 - 项目类别:
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