Opsineering: Engineering Novel ChannelRhodospins for Optogenetics Applications

Opsineering：为光遗传学应用设计新型通道Rhodospins

基本信息

批准号：
9125904
负责人：
Claire Nicole Bedbrook
金额：
$ 3.72万
依托单位：
CALIFORNIA INSTITUTE OF TECHNOLOGY
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-09-01 至 2017-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9125904
关键词：
Address Animals Autistic Disorder Biological Biological Assay Brain Brain region Cations Cells Characteristics Chemical Stimulation Chimera organism Collaborations Color Development Diagnosis Directed Molecular Evolution Drug abuse Electric Stimulation Electrophysiology (science)Engineering Evolution Family Foundations Gated Ion Channel Genetic Recombination Goals Health Homologous Gene Ions Kinetics Laboratories Length Libraries Life Light Literature Mammalian Cell Membrane Mental Depression Mental disorders Methods Modeling Mutate Mutation Neurobiology Neurons Neurosciences Opsin Organism Parkinson Disease Pathway interactions Permeability Photophobia Population Heterogeneity Probability Property Protein Engineering Proteins Research Research Training Schizophrenia Specific qualifier value Specificity Speed Stimulus Structure System Techniques Time Transgenic Organisms Variant Work addiction base brain circuitry cell type desensitization design improved light gated member microbial millisecond mutant nervous system disorder neuronal circuitry next generation novel optogenetics protein function screening single molecule success tool

项目摘要

DESCRIPTION (provided by applicant): Optogenetics is genetically encoded, optically induced, control of cells through transgenic expression of microbial opsins in mammalian neurons. When these opsins are expressed in a cell-type specific manner and light activated, they provide temporally and spatially separated stimulation of independent hyperpolarizing and depolarizing channels in neurons in living animals. Channelrhodopsins (ChRs) are the microbial opsins used in optogenetics to trigger light induced depolarization. ChRs are light-gated ion channels that operate on the order of milliseconds, a time scale relevant for neuronal activation, and can be expressed in the membrane of distinct cell types with high temporal precision in well-defined brain regions. This contrasts with the poor temporal dynamics or lack of specificity of chemical or electrical stimulation methods. However, the optogenetics tools currently available for neuronal circuit interrogation are limited based on expression, light-wavelength activation, kinetics and ion specificity. Our proposed project addresses these limitations through protein engineering. Protein engineering through directed evolution and structure-guided recombination are well-established methods for modifying and optimizing proteins for desired functions. Current literature and preliminary collaborative work between the Gradinaru and Arnold labs at Caltech indicate that channelrhodopsins are amenable to functionally useful laboratory evolution and manipulation. This work will be focused toward engineering improved channelrhodopsins for use as biological tools in optogenetics. The aim is to engineer channelrhodopsins for optimal ion selectivity, kinetics, reversibility, and shifted light excitatio wavelengths. These new channel proteins will have applications in probing the brain's circuitry to better understand and model healthy and non-healthy brain function as a foundation for controlling and diagnosing neurological disorders such as addiction, depression and Parkinson's disease.

描述（由申请人提供）：光遗传学是通过在哺乳动物神经元中微生物视蛋白的转基因表达来对细胞进行遗传编码、光诱导和控制。当这些视蛋白以细胞类型特异性方式表达并被光激活时，它们对活体动物神经元中独立的超极化和去极化通道提供时间和空间上分离的刺激。视紫红质通道蛋白 (ChRs) 是光遗传学中用于触发光诱导去极化的微生物视蛋白。 ChR 是光门控离子通道，其运行时间为毫秒级（与神经元激活相关的时间尺度），并且可以在明确的大脑区域中以高时间精度在不同细胞类型的膜中表达。这与化学或电刺激方法的时间动态性差或缺乏特异性形成鲜明对比。然而，目前可用于神经元回路询问的光遗传学工具由于表达、光波长激活、动力学和离子特异性而受到限制。我们提出的项目通过蛋白质工程解决了这些限制。通过定向进化和结构引导重组的蛋白质工程是修饰和优化蛋白质以获得所需功能的成熟方法。目前的文献以及加州理工学院 Gradinaru 和 Arnold 实验室之间的初步合作工作表明，通道视紫红质适合功能上有用的实验室进化和操作。这项工作将集中于工程改良视紫红质通道，用作光遗传学中的生物工具。目的是设计视紫红质通道以获得最佳的离子选择性、动力学、可逆性和移动的光激发波长。这些新的通道蛋白将应用于探测大脑的电路，以更好地理解和模拟健康和不健康的大脑功能，作为控制和诊断神经系统疾病（如成瘾、抑郁症和帕金森病）的基础。

项目成果

期刊论文数量（7）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

AUGMENTING PROTEIN NETWORK EMBEDDINGS

增强蛋白质网络嵌入

DOI：
发表时间：
2024-09-13
期刊：
影响因子：
0
作者：
通讯作者：

Machine learning to design integral membrane channelrhodopsins for efficient eukaryotic expression and plasma membrane localization.

机器学习设计完整的膜通道视紫红质，以实现有效的真核表达和质膜定位。

DOI：
发表时间：
2017-10
期刊：
影响因子：
4.3
作者：
Bedbrook, Claire N;Yang, Kevin K;Rice, Austin J;Gradinaru, Viviana;Arnold, Frances H
通讯作者：
Arnold, Frances H

Directed Evolution of a Bright Near-Infrared Fluorescent Rhodopsin Using a Synthetic Chromophore.

使用合成发色团定向进化明亮的近红外荧光视紫红质。

DOI：
10.1016/j.chembiol.2017.02.008
发表时间：
2017-03-16
期刊：
Cell chemical biology
影响因子：
8.6
作者：
Herwig L;Rice AJ;Bedbrook CN;Zhang RK;Lignell A;Cahn JKB;Renata H;Dodani SC;Cho I;Cai L;Gradinaru V;Arnold FH
通讯作者：
Arnold FH

Structure-guided SCHEMA recombination of distantly related beta-lactamases.

远缘相关β-内酰胺酶的结构引导模式重组。