Real time optimization of electron-based fragmentation for middle and top-down proteomics in mass spectrometry

质谱中中自上而下蛋白质组学基于电子的碎片实时优化

• 批准号: 10081127
• 负责人: Valery G. Voinov
• 金额: $ 21.28万
• 依托单位: E-MSION, INC.
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-01 至 2021-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10081127
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The identification and quantification of biological macromolecules remains challenging despite major advances in the speed, resolution and mass accuracy of modern mass spectrometers. A key weakness with current instrumentation lies in the methods used to induce fragmentation. The reliance in particular on collision-induced dissociation (CID) has limited such analyses to bottom-up workflows of trypsin-digested peptides of 10-30 residues. At e-MSion, we have developed an efficient electron-fragmentation technology called ExD for large proteins and are now co-marketed our ExD Option with Agilent, and soon will be with Thermo and Waters instruments. What has really captured the interest of the biopharma and top-down communities in the past year is the exceptional sequence coverage of native proteins we obtain with the same ExD cell. The resulting spectra are less congested than those obtained with currently available ETD/UVPD/CID fragmentation methodologies. We have shown that our technology works faster and gives cleaner spectra with more complete dissociation with larger macromolecular protein complexes than has ever been possible before, while still preserving labile post translational modifications. In addition, fragmentation with higher energy electrons can be used to provide complementary data to improve protein and glycan identification. The challenge now has become how to optimally collect and process these data to maximize the utility of ExD fragmentation. Last summer, Xilinx released its Versal Adaptive Compute Acceleration Platform (ACAP), a massively parallel processor with 50 billion transistors targeted to transform digital signal processing, handling of big data and artificial intelligence. This ACAP technology has already accelerated Illumina DNA sequence assembly by 90-fold. Our feasibility question asks how to effectively harness this new highly parallelized technology to preprocess complex top-down mass spectra on- the-fly. This will allow us to actively optimize data acquisition by enabling adaptive operation of the ExD cell and mass spectrometer. The objective is to maximize both fragmentation and dissociation of native proteins, enabling faster and comprehensive characterization of challenging proteoforms important to the biopharmaceutical industry and biomedical researchers. Success will offer an extremely fast, cost-effective solution to characterize complexes of macromolecules under native conditions with increased accuracy, speed, and fewer misidentifications. Our ExD technology with the Versal ACAP can be both retrofitted into existing mass spectrometers as well as being available in new generations of mass spectrometers at a price below other less-effective alternative fragmentation technologies like ETD and UVPD. Thus, it will provide new abilities for many NIH investigators to advance basic research, probe disease mechanisms and permit more sophisticated searches for both diagnostic and therapeutic biomarkers.

尽管存在重大问题，生物大分子的鉴定和定量仍然具有挑战性 现代质谱仪在速度、分辨率和质量精度方面取得了进步。一个关键弱点是 当前的仪器在于用于诱导碎片的方法。尤其是依赖于 碰撞诱导解离 (CID) 将此类分析限制为胰蛋白酶消化的自下而上的工作流程 10-30个残基的肽。在 e-MSion，我们开发了一种高效的电子碎片技术 用于大蛋白质的 ExD，现在与安捷伦共同销售我们的 ExD 选项，很快将与 Thermo 和 Waters 仪器。是什么真正引起了生物制药公司和自上而下的兴趣 去年的社区是我们用相同的方法获得的天然蛋白质的特殊序列覆盖率 ExD 细胞。所得光谱比目前可用的光谱更不拥挤 ETD/UVPD/CID 碎片方法。我们已经证明，我们的技术运行速度更快，并且能够提供 与更大的大分子蛋白质复合物相比，光谱更清晰，解离更完全 以前曾经有过这种情况，同时仍然保留不稳定的翻译后修饰。此外， 具有更高能量电子的碎片可用于提供补充数据以改进蛋白质 和聚糖鉴定。现在的挑战是如何以最佳方式收集和处理这些数据 最大化 ExD 碎片的效用。去年夏天，Xilinx 发布了 Versal 自适应计算 加速平台 (ACAP)，一个具有 500 亿个晶体管的大规模并行处理器，旨在 转变数字信号处理、大数据处理和人工智能。该 ACAP 技术具有 已经将 Illumina DNA 序列组装速度加快了 90 倍。我们的可行性问题询问如何 有效地利用这种新的高度并行技术来预处理复杂的自上而下的质谱图 苍蝇。这将使我们能够通过启用 ExD 单元的自适应操作来主动优化数据采集 和质谱仪。目标是最大化天然蛋白质的断裂和解离， 能够更快、更全面地表征对重要的具有挑战性的蛋白质形式 生物制药行业和生物医学研究人员。 成功将提供一种极其快速、具有成本效益的解决方案来表征复杂的 在天然条件下识别大分子，具有更高的准确性、速度和更少的错误识别。我们的 ExD 技术与 Versal ACAP 可以改装为现有的质谱仪以及 可用于新一代质谱仪，价格低于其他效率较低的替代品 ETD 和 UVPD 等碎片技术。因此，它将为许多 NIH 提供新的能力 研究人员推进基础研究、探索疾病机制并允许更复杂的搜索 用于诊断和治疗生物标志物。