Rational design and functionalization of circular tandem repeat proteins

环状串联重复蛋白的合理设计和功能化

• 批准号: 9897572
• 负责人: Philip Bradley
• 金额: $ 34.54万
• 依托单位: FRED HUTCHINSON CANCER RESEARCH CENTER
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-05-01 至 2022-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9897572
• 关键词: Adaptor Signaling Protein; Algorithm Design; Algorithms; Architecture; Avidity; Binding; Biochemical; Biological Process; Biophysics; Caliber; Cell Therapy; Cell surface; Cells; Cellular Assay; Clinical; Collaborations; Communities; Complex; Computational algorithm; Computing Methodologies; Development; Diagnostic Reagent; Dimensions; Elements; Evolution; Extracellular Domain; Family; Gene Proteins; Geometry; Goals; Handedness; Hematopoietic stem cells; Homo; IL3 Gene; Interleukin-2; Length; Location; Methods; Molecular; Molecular Conformation; Mutation; Nature; Organism; Output; Peptides; Property; Protein Array; Protein Engineering; Protein Region; Proteins; Reagent; Recording of previous events; Reporting; Research; Sampling; Scaffolding Protein; Side; Signal Transduction; Site; Source; Specificity; Speed; Structure; Surface; System; T-Lymphocyte; Tandem Repeat Sequences; Tertiary Protein Structure; Testing; Therapeutic; Validation; Work; cellular development; cytokine; design; design and construction; gene synthesis; improved; member; novel; novel strategies; protein expression; protein folding; rational function; scaffold; src Homology Region 2 Domain; tool

PROJECT SUMMARY/ABSTRACT Natural evolution has produced a stunningly diverse array of proteins that perform an equally diverse set of molecular functions in living organisms. These proteins—which constitute the primary raw material from which we might seek to develop new protein reagents—have been honed over the course of their mutational history to meet specific functional challenges. As a result, turning them to new functions by rational means often proves problematic: their expression and/or stability are compromised by our reengineering attempts, and their relic functionality is at odds with our intended use. De novo protein design, which uses sophisticated computer algorithms to identify stable sequence:structure pairings without relying on native templates, can create protein folds never before seen in Nature, and thus offers an alternative source of protein scaffolds for functionalization. We recently reported the development of new algorithms for de novo design of a particular class of proteins— circular tandem repeat proteins or cTRPs—whose modular, self-reinforcing symmetrical architecture offers advantages that include high stability, tunable geometry, and switchable oligomeric state. We hypothesize that de novo designed proteins in general, and these designed cTRPs in particular, will prove to be a valuable source of protein scaffolds for downstream application. Our aims in this proposal are first, to further develop our algorithms in order to design and experimentally validate a diverse set of cTRP scaffolds of varied size and topology; and second, in collaboration with clinical colleagues here at the Hutchinson Center, to evaluate these designs as scaffolds for presentation of functional domains with precisely controlled symmetry and geometry. Our collaborators will test these designed constructs in cellular assays with the goal of speeding the development of cellular therapies. Successful completion of this research will lead to (1) improved protein design algorithms that have been rigorously validated across a range of topologies and are available to the research community; (2) a family of stable and robust protein scaffolds for downstream functionalization, all of whose members have been structurally and biophysically characterized; (3) a set of useful protein reagents for biomedical applications.

项目摘要/摘要 自然进化产生了令人惊叹的各种蛋白质，它们执行了同样多样的蛋白质 生物中的分子功能。这些蛋白质 - 构成主要原材料 我们可能会寻求开发新的蛋白质试剂 - 在其突变史的过程中受到尊重 应对特定的功能挑战。结果，经常通过理性手段将它们转向新功能 证明有问题的是：我们的表达和/或稳定性因我们的重新设计尝试而损害了 遗物功能与我们的预期用途不符。从头蛋白质设计，使用复杂的计算机 识别稳定序列的算法：结构配对而不依赖天然模板，可以创建蛋白质 褶皱在自然界中从未见过，因此提供了用于功能化的蛋白质支架的替代来源。 我们最近报道了开发针对特定类别蛋白质的从头设计的新算法 - 圆形串联重复蛋白或CTRP-其模块化，自我强化对称体系结构提供 包括高稳定性，可调几何形状和可切换寡聚状态的优点。我们假设这一点 从头设计的蛋白质一般设计，尤其是这些蛋白质，这些蛋白质将被证明是有价值的 用于下游应用的蛋白质支架的来源。我们在该提案中的目标是进一步发展 我们的算法是为了设计和实验验证各种大小的CTRP支架和 拓扑；其次，在哈钦森中心与临床同事合作以评估这些 设计作为脚手架，以呈现具有精确控制的对称性和几何形状的功能域。 我们的合作者将测试这些在蜂窝测定中的设计结构，目的是加速 细胞疗法的发展。成功完成这项研究将导致（1）改善蛋白质 设计算法已在一系列拓扑中进行了严格验证，可用于 研究社区； （2）一个稳定且健壮的蛋白质支架的家族，用于下游功能，所有 其成员在结构和生物物理上的表征； （3）一组有用的蛋白质试剂 生物医学应用。