一位大咖用Octet发表了20篇CNS！
ai药物在近几年发展迅速，其中，蛋白质从头设计无疑是ai设计药物中的重头戏与热点。它不仅可以针对不可成药靶点的药物和自然中不存在的蛋白进行设计，而且大大缩短了药物研发的周期。
华盛顿大学医学院的david baker教授是该领域的强者。当然，ai再牛，也需要用实验手段去表征设计的蛋白。比如，使用octet® 非标记分子互作系统检测设计的蛋白与靶点的亲和力。目前，david baker课题组已使用octet® 检测蛋白亲和力发表了近40篇文章，其中cns文章就有近20篇！(文章列表见文末)
这里，陈老师就介绍一下他今年发表的几篇cns文章。
nature
de novo design of protein structure and function with rf diffusion[1]
本文结合扩散模型（rf diffusion）和用深度学习算法，实现了从头设计合成蛋白质的目标，设计成功率高两个数量级。针对已知的五个靶标，只需要不到100个候选即可达到nm级别的亲和力（全部用bli进行检测）。同时，研究人员还设计了一种蛋白与其底物（流感血凝素）的复合物，并使用冷冻电镜解析了其结构。结果显示，冷冻电镜解析的结构与设计的模型几乎一模一样，从而证明了该模型的准确性。为了进一步验证生成的蛋白是否具有结合活性，研究人员继续使用octet®对这两种蛋白的结合进行验证。
图1.(a-c) 从头设计靶标蛋白的结合蛋白，针对五个靶标，从头设计结合蛋白，bli 响应信号值≥阳性对照1/2为候选;（b）rf diffusion成功率高出两个数量级; (d) 亲和力最高的结合物，结合kd为28nm; (e-h) 结构学验证
研究团队表示，rf diffusion是对目前蛋白质设计方法的一次综合改进，能够产生总长度达600个氨基酸残基的结构，其复杂性和准确度均比之前更高。研究团队还表示，对该方法的进一步改进或能设计出复杂程度更高的全新蛋白质。
nature
de novo design of modular peptite-binding proteins by superhelical matching
针对内在无序区的重复性蛋白和多肽，本文开发了一种从头设计蛋白的通用性方法，能以大约 20% 的成功率高效设计出多肽结合蛋白。这些蛋白具备低至皮摩（pm）级别的高亲和力、高特异性、高热稳定性以及高精度。而且还证实了该方法可以靶向更广阔的非重复性多肽区域、以及可以延伸至人源蛋白的复杂网络。
审稿人评价称：“表征非常详尽，亲和力高达纳摩尔至皮摩尔范围，并且具有很高的特异性。设计与结构的匹配也表明核心方法是合理的。”
这种高亲和力、高特异性就是通过octet® 来进行检测的。
图2.将生物素化的目标肽加载到生物传感器上，并和设计的binder进行结合解离，红色矩形框表示相匹配的结合；可见，设计的binder与目标多肽的良好特异性
science
top-down design of protein architectures with reinforcement learning
该研究开发了一种基于强化学习的蛋白质设计软件，并证明了它有能力创造有功能的蛋白质。这项工作用于设计高度稳定和多功能的多聚蛋白笼组装结构和纳米蛋白颗粒，开启了蛋白质设计的新时代。该技术对癌症治疗、再生医学、强效疫苗和可生物降解日用品都有积极影响，文章同样使用octet® 检测了设计的多聚蛋白颗粒与已知表位抗体的结合，以进一步验证其结构的正确性。
图3.octet® 亲和力测定发现设计的单体蛋白聚合颗粒可以与识别不同表位抗体结合，表明单体蛋白在聚合颗粒上的构象依旧完整。
octet® 为什么如此受欢迎呢？
因为octet® 用于亲和力验证的优点在于
非标记direct binding是趋势，不需要标记和信号放大，可以更好的保持反应物的活性
快速测定亲和力，提供结合速率常数和解离速率常数更加定量化地表征分子互作
无洗涤步骤，可测弱亲和力（解离快）
写入了美国药典，文章多，认可度广
万金油技术，可以用与检测dna，小分子，蛋白质等各种生物分子
操作简便，耗材及维护成本低
尽管经过不同的ai算法优化，binder设计的成功率已有所提高，但目前设计出高亲和力的binder仍然具有相当大的难度。对ppi（蛋白质相互作用）的建模和理解仍然需要进行大量的工作。octet® 可以真正实现高通量、快速的表征和检测，这极大地加速了科研进程，实现预测和验证的有机结合。
-参考文献
[1] de novo design of protein structure and function with rfdiffusion
jl watson, d juergens, nr bennett, bl trippe, j yim… - nature, 2023 - nature.com
[2] de novo design of modular peptide-binding proteins by superhelical matching.nature,2023
[3] top-down design of protein architectures with reinforcement learning.
sciencevol. 380, no. 6642
[4] massively parallel de novo protein design for targeted therapeutics
…, x huang, r jin, ia wilson, dh fuller, d baker - nature, 2017 - nature.com
[5] optimization of affinity, specificity and function of designed influenza inhibitors using deep sequencing
…, h kamisetty, p blair, ia wilson, d baker - nature …, 2012 - nature.com
[6] de novo design of picomolar sars-cov-2 miniprotein inhibitors
…, l stewart, ms diamond, d veesler, d baker - science, 2020 - science.org
[7] de novo design of bioactive protein switches
…, je dueber, wrp novak, h el-samad, d baker - nature, 2019 - nature.com
[8] high-throughput characterization of protein–protein interactions by reprogramming yeast mating
d younger, s berger, d baker… - proceedings of the …, 2017 - national acad sciences
[9] a potent anti-malarial human monoclonal antibody targets circumsporozoite protein minor repeats and neutralizes sporozoites in the liver
…, r vistein, c barillas-mury, r amino, d baker… - immunity, 2020 - elsevier
[10] quadrivalent influenza nanoparticle vaccines induce broad protection
…, mc crank, l stewart, kk lee, m guttman, d baker… - nature, 2021 - nature.com
[11] transferrin receptor targeting by de novo sheet extension
…, de ingber, j abraham, d baker - proceedings of the …, 2021 - national acad sciences
[12] receptor subtype discrimination using extensive shape complementary designed interfaces
…, cj kuo, kc garcia, d baker - nature structural & …, 2019 - nature.com
[13] ultrapotent miniproteins targeting the sars-cov-2 receptor-binding domain protect against infection and disease
…, r ravichandran, l carter, l stewart, d baker… - cell host & microbe, 2021 - elsevier
[14] targeting hiv env immunogens to b cell follicles in nonhuman primates through immune complex or protein nanoparticle formulations
…, g alter, wr schief, s crotty, np king, d baker… - npj vaccines, 2020 - nature.com
[15] de novo design of potent and selective mimics of il-2 and il-15
…, gjl bernardes, m dougan, kc garcia, d baker - nature, 2019 - nature.com
[16] computational design of proteins targeting the conserved stem region of influenza hemagglutinin
…, c dreyfus, je corn, em strauch, ia wilson, d baker - science, 2011 - science.org
[17] reconfigurable asymmetric protein assemblies through implicit negative design
…, j decarreau, hm morris, a kang, ak bera, d baker - science, 2022 - science.org
[18] structural and functional evaluation of de novo-designed, two-component nanoparticle carriers for hiv env trimer immunogens
…, jp moore, rw sanders, np king, d baker… - plos …, 2020 - journals.plos.org
[19] polyclonal antibody responses to hiv env immunogens resolved using cryoem
…, rf rocha, zt berndsen, d baker… - nature …, 2021 - nature.com
[20] induction of potent neutralizing antibody responses by a designed protein nanoparticle vaccine for respiratory syncytial virus
…, kk lee, d veesler, ce correnti, lj stewart, d baker… - cell, 2019 - elsevier
[21] designed protein logic to target cells with precise combinations of surface antigens
…, a nguyen, s pun, ce correnti, sr riddell, d baker - science, 2020 - science.org
[22] de novo design of tyrosine and serine kinase-driven protein switches
…, vh wysocki, h el-samad, d baker - nature structural & …, 2021 - nature.com
[23] computational design of trimeric influenza-neutralizing proteins targeting the hemagglutinin receptor binding site
…, ab ward, p yager, dh fuller, ia wilson, d baker - nature …, 2017 - nature.com
[24] scaffolding protein functional sites using deep learning
…, f dimaio, b correia, s ovchinnikov, d baker - science, 2022 - science.org
[25] m*lent designed proteins neutralize sars-cov-2 variants of concern and confer protection against infection in mice
…, bs freedman, jd bloom, h ruohola-baker… - science translational …, 2022 - science.org
[26] a computationally designed hemagglutinin stem-binding protein provides in vivo protection from influenza independent of a host immune response
…, ia wilson, a dagley, df smee, d baker… - plos …, 2016 - journals.plos.org
[27] computational design of a synthetic pd-1 agonist
…, f dimaio, kv tarbell, d baker - proceedings of the …, 2021 - national acad sciences
[28] first critical repressive h3k27me3 marks in embryonic stem cells identified using designed protein inhibitor
…, sh orkin, d baker, h ruohola-baker - proceedings of the …, 2017 - national acad sciences
[29] designed proteins assemble antibodies into modular nanocages
…, h ruohola-baker, j mathieu, d veesler, d baker - science, 2021 - science.org
[30] designed protein logic to target cells with precise combinations of surface antigens
…, a nguyen, s pun, ce correnti, sr riddell, d baker - science, 2020 - science.org
[31] de novo design of tyrosine and serine kinase-driven protein switches
…, vh wysocki, h el-samad, d baker - nature structural & …, 2021 - nature.com
[32] computational design of trimeric influenza-neutralizing proteins targeting the hemagglutinin receptor binding site
…, ab ward, p yager, dh fuller, ia wilson, d baker - nature …, 2017 - nature.com
[33] scaffolding protein functional sites using deep learning
…, f dimaio, b correia, s ovchinnikov, d baker - science, 2022 - science.org
[34] m*lent designed proteins neutralize sars-cov-2 variants of concern and confer protection against infection in mice
…, bs freedman, jd bloom, h ruohola-baker… - science translational …, 2022 - science.org
[35] a computationally designed hemagglutinin stem-binding protein provides in vivo protection from influenza independent of a host immune response
…, ia wilson, a dagley, df smee, d baker… - plos …, 2016 - journals.plos.org
[36] computational design of a synthetic pd-1 agonist
…, f dimaio, kv tarbell, d baker - proceedings of the …, 2021 - national acad sciences
[37] first critical repressive h3k27me3 marks in embryonic stem cells identified using designed protein inhibitor
…, sh orkin, d baker, h ruohola-baker - proceedings of the …, 2017 - national acad sciences
[38] designed proteins assemble antibodies into modular nanocages
…, h ruohola-baker, j mathieu, d veesler, d baker - science, 2021 - science.org
[39] computational design of a ph-sensitive igg binding protein
…, sj fleishman, d baker - proceedings of the …, 2014 - national acad sciences
[40] design of protein-binding proteins from the target structure alone
…, s bernard, l stewart, ia wilson, h ruohola-baker… - nature, 2022 - nature.com
[41] de novo design of modular and tunable protein biosensors
…, j wi, hj hong, l stewart, bh oh, d baker - nature, 2021 - nature.com
科研用乳酸林格氏液的使用方法
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一位大咖用Octet发表了20篇CNS！
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华盛顿大学医学院的david baker教授是该领域的强者。当然，ai再牛，也需要用实验手段去表征设计的蛋白。比如，使用octet® 非标记分子互作系统检测设计的蛋白与靶点的亲和力。目前，david baker课题组已使用octet® 检测蛋白亲和力发表了近40篇文章，其中cns文章就有近20篇！(文章列表见文末)
这里，陈老师就介绍一下他今年发表的几篇cns文章。
nature
de novo design of protein structure and function with rf diffusion[1]
本文结合扩散模型（rf diffusion）和用深度学习算法，实现了从头设计合成蛋白质的目标，设计成功率高两个数量级。针对已知的五个靶标，只需要不到100个候选即可达到nm级别的亲和力（全部用bli进行检测）。同时，研究人员还设计了一种蛋白与其底物（流感血凝素）的复合物，并使用冷冻电镜解析了其结构。结果显示，冷冻电镜解析的结构与设计的模型几乎一模一样，从而证明了该模型的准确性。为了进一步验证生成的蛋白是否具有结合活性，研究人员继续使用octet®对这两种蛋白的结合进行验证。
图1.(a-c) 从头设计靶标蛋白的结合蛋白，针对五个靶标，从头设计结合蛋白，bli 响应信号值≥阳性对照1/2为候选;（b）rf diffusion成功率高出两个数量级; (d) 亲和力最高的结合物，结合kd为28nm; (e-h) 结构学验证
研究团队表示，rf diffusion是对目前蛋白质设计方法的一次综合改进，能够产生总长度达600个氨基酸残基的结构，其复杂性和准确度均比之前更高。研究团队还表示，对该方法的进一步改进或能设计出复杂程度更高的全新蛋白质。
nature
de novo design of modular peptite-binding proteins by superhelical matching
针对内在无序区的重复性蛋白和多肽，本文开发了一种从头设计蛋白的通用性方法，能以大约 20% 的成功率高效设计出多肽结合蛋白。这些蛋白具备低至皮摩（pm）级别的高亲和力、高特异性、高热稳定性以及高精度。而且还证实了该方法可以靶向更广阔的非重复性多肽区域、以及可以延伸至人源蛋白的复杂网络。
审稿人评价称：“表征非常详尽，亲和力高达纳摩尔至皮摩尔范围，并且具有很高的特异性。设计与结构的匹配也表明核心方法是合理的。”
这种高亲和力、高特异性就是通过octet® 来进行检测的。
图2.将生物素化的目标肽加载到生物传感器上，并和设计的binder进行结合解离，红色矩形框表示相匹配的结合；可见，设计的binder与目标多肽的良好特异性
science
top-down design of protein architectures with reinforcement learning
该研究开发了一种基于强化学习的蛋白质设计软件，并证明了它有能力创造有功能的蛋白质。这项工作用于设计高度稳定和多功能的多聚蛋白笼组装结构和纳米蛋白颗粒，开启了蛋白质设计的新时代。该技术对癌症治疗、再生医学、强效疫苗和可生物降解日用品都有积极影响，文章同样使用octet® 检测了设计的多聚蛋白颗粒与已知表位抗体的结合，以进一步验证其结构的正确性。
图3.octet® 亲和力测定发现设计的单体蛋白聚合颗粒可以与识别不同表位抗体结合，表明单体蛋白在聚合颗粒上的构象依旧完整。
octet® 为什么如此受欢迎呢？
因为octet® 用于亲和力验证的优点在于
非标记direct binding是趋势，不需要标记和信号放大，可以更好的保持反应物的活性
快速测定亲和力，提供结合速率常数和解离速率常数更加定量化地表征分子互作
无洗涤步骤，可测弱亲和力（解离快）
写入了美国药典，文章多，认可度广
万金油技术，可以用与检测dna，小分子，蛋白质等各种生物分子
操作简便，耗材及维护成本低
尽管经过不同的ai算法优化，binder设计的成功率已有所提高，但目前设计出高亲和力的binder仍然具有相当大的难度。对ppi（蛋白质相互作用）的建模和理解仍然需要进行大量的工作。octet® 可以真正实现高通量、快速的表征和检测，这极大地加速了科研进程，实现预测和验证的有机结合。
-参考文献
[1] de novo design of protein structure and function with rfdiffusion
jl watson, d juergens, nr bennett, bl trippe, j yim… - nature, 2023 - nature.com
[2] de novo design of modular peptide-binding proteins by superhelical matching.nature,2023
[3] top-down design of protein architectures with reinforcement learning.
sciencevol. 380, no. 6642
[4] massively parallel de novo protein design for targeted therapeutics
…, x huang, r jin, ia wilson, dh fuller, d baker - nature, 2017 - nature.com
[5] optimization of affinity, specificity and function of designed influenza inhibitors using deep sequencing
…, h kamisetty, p blair, ia wilson, d baker - nature …, 2012 - nature.com
[6] de novo design of picomolar sars-cov-2 miniprotein inhibitors
…, l stewart, ms diamond, d veesler, d baker - science, 2020 - science.org
[7] de novo design of bioactive protein switches
…, je dueber, wrp novak, h el-samad, d baker - nature, 2019 - nature.com
[8] high-throughput characterization of protein–protein interactions by reprogramming yeast mating
d younger, s berger, d baker… - proceedings of the …, 2017 - national acad sciences
[9] a potent anti-malarial human monoclonal antibody targets circumsporozoite protein minor repeats and neutralizes sporozoites in the liver
…, r vistein, c barillas-mury, r amino, d baker… - immunity, 2020 - elsevier
[10] quadrivalent influenza nanoparticle vaccines induce broad protection
…, mc crank, l stewart, kk lee, m guttman, d baker… - nature, 2021 - nature.com
[11] transferrin receptor targeting by de novo sheet extension
…, de ingber, j abraham, d baker - proceedings of the …, 2021 - national acad sciences
[12] receptor subtype discrimination using extensive shape complementary designed interfaces
…, cj kuo, kc garcia, d baker - nature structural & …, 2019 - nature.com
[13] ultrapotent miniproteins targeting the sars-cov-2 receptor-binding domain protect against infection and disease
…, r ravichandran, l carter, l stewart, d baker… - cell host & microbe, 2021 - elsevier
[14] targeting hiv env immunogens to b cell follicles in nonhuman primates through immune complex or protein nanoparticle formulations
…, g alter, wr schief, s crotty, np king, d baker… - npj vaccines, 2020 - nature.com
[15] de novo design of potent and selective mimics of il-2 and il-15
…, gjl bernardes, m dougan, kc garcia, d baker - nature, 2019 - nature.com
[16] computational design of proteins targeting the conserved stem region of influenza hemagglutinin
…, c dreyfus, je corn, em strauch, ia wilson, d baker - science, 2011 - science.org
[17] reconfigurable asymmetric protein assemblies through implicit negative design
…, j decarreau, hm morris, a kang, ak bera, d baker - science, 2022 - science.org
[18] structural and functional evaluation of de novo-designed, two-component nanoparticle carriers for hiv env trimer immunogens
…, jp moore, rw sanders, np king, d baker… - plos …, 2020 - journals.plos.org
[19] polyclonal antibody responses to hiv env immunogens resolved using cryoem
…, rf rocha, zt berndsen, d baker… - nature …, 2021 - nature.com
[20] induction of potent neutralizing antibody responses by a designed protein nanoparticle vaccine for respiratory syncytial virus
…, kk lee, d veesler, ce correnti, lj stewart, d baker… - cell, 2019 - elsevier
[21] designed protein logic to target cells with precise combinations of surface antigens
…, a nguyen, s pun, ce correnti, sr riddell, d baker - science, 2020 - science.org
[22] de novo design of tyrosine and serine kinase-driven protein switches
…, vh wysocki, h el-samad, d baker - nature structural & …, 2021 - nature.com
[23] computational design of trimeric influenza-neutralizing proteins targeting the hemagglutinin receptor binding site
…, ab ward, p yager, dh fuller, ia wilson, d baker - nature …, 2017 - nature.com
[24] scaffolding protein functional sites using deep learning
…, f dimaio, b correia, s ovchinnikov, d baker - science, 2022 - science.org
[25] m*lent designed proteins neutralize sars-cov-2 variants of concern and confer protection against infection in mice
…, bs freedman, jd bloom, h ruohola-baker… - science translational …, 2022 - science.org
[26] a computationally designed hemagglutinin stem-binding protein provides in vivo protection from influenza independent of a host immune response
…, ia wilson, a dagley, df smee, d baker… - plos …, 2016 - journals.plos.org
[27] computational design of a synthetic pd-1 agonist
…, f dimaio, kv tarbell, d baker - proceedings of the …, 2021 - national acad sciences
[28] first critical repressive h3k27me3 marks in embryonic stem cells identified using designed protein inhibitor
…, sh orkin, d baker, h ruohola-baker - proceedings of the …, 2017 - national acad sciences
[29] designed proteins assemble antibodies into modular nanocages
…, h ruohola-baker, j mathieu, d veesler, d baker - science, 2021 - science.org
[30] designed protein logic to target cells with precise combinations of surface antigens
…, a nguyen, s pun, ce correnti, sr riddell, d baker - science, 2020 - science.org
[31] de novo design of tyrosine and serine kinase-driven protein switches
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