Analysis of the mechanism of protein secretion through the Sec machinery and exploitation as a polypeptide sequencing device

通过 Sec 机制分析蛋白质分泌机制并开发为多肽测序装置

• 批准号: 2885488
• 金额: --
• 依托单位: University of Bristol
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2023
• 资助国家: 英国
• 起止时间: 2023 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2885488
• 关键词: Analysis; mechanism; protein; secretion; through

Transport of proteins across membranes is a fundamental biological process essential for protein secretion andorganelle biogenesis1,2. This CASE project concerns the bacterial system, wherein protein transport from the cytosolacross the inner-membrane is usually achieved when the SecYEG protein-channel complex engages the cytosolic motorATPase SecA (secretion). Great strides have been made towards understanding the mechanism of protein-translocation:firstly, through the determination of the structures of the protein-channel and the motor components 3,4, and secondly,through the development of accurate and high-resolution assays for protein transport. We have developed such anassay, based on a split luciferase system, for both mitochondrial import and bacterial secretion5,6. The project willcontinue with the exploitation of this technology, together with a wide range of biochemical and biophysicalapproaches, towards the determination of the underlying molecular basis for protein transport. In parallel, your CASE project will be partnered with Oxford Nanopore Technologies (ONT) to explore the prospect ofexploiting the channel as a polypeptide sequencer. This would be achieved in the spirit of ONT technology developed forDNA sequencing - by monitoring variable conductance as different nucleotides of a single polymer pass through a porein the membrane. Currently, peptide sequencing is very challenging, time consuming and expensive, so if this can besimplified, and adapted for biological samples then the implications for analytical biochemistry and cell biology research,diagnostics, forensics etc. would be game changing. Early indications are very promising as we know that positivecharges (lysines and particularly arginines) and bulky residues struggle to make it through the channel and slowtransport considerably5. Therefore, different residues must have distinct interactions with the SecY-channel and mightalso elicit a measurable difference in conductance, and thereby generate interpretable signatures for different residuesrequired for sequencing. ONT is a globally successful company with a portfolio of technological innovations. Therefore,the partnership will bring together expertise of the Sec machinery together with sequencing know-how and platforms tocreate an ideal environment for the success of your project, for training and for experience of academic and biotechenvironments and teamwork.

蛋白质跨膜的运输是蛋白质分泌必不可少的基本生物学过程Andorganelle生物发生1,2。该病例项目涉及细菌系统，其中通常从细胞质酸转运的蛋白转运时，当Secyeg蛋白通道复合物与细胞质电动型SECA（分泌）接合时，通常会实现内膜。在理解蛋白质转换的机理方面取得了长足的进步：首先，通过确定蛋白质通道的结构和运动成分3,4，其次是通过开发准确和高分辨率的蛋白质转运测定法。我们已经基于分裂荧光素酶系统开发了这种Anassay，用于线粒体进口和细菌分泌5,6。该项目将随着该技术的利用以及广泛的生化和生物物理攻击，以确定蛋白质转运的基本分子基础。同时，您的案例项目将与牛津纳米孔技术（ONT）合作，以探索作为多肽序列验证通道的前景。这将以ONT技术的精神开发的fordna测序来实现 - 通过监测可变电导，因为单个聚合物的不同核苷酸通过膜通过孔洞。当前，肽测序非常具有挑战性，耗时且昂贵，因此，如果可以对其进行简化并适合生物样品，那么对分析生物化学和细胞生物学研究，诊断，取证等的影响将在改变游戏。早期的迹象非常有前途，因为我们知道，阳性疗法（赖氨酸，尤其是精氨酸）和庞大的残留物难以通过通道和慢速转移5。因此，不同的残基必须与SECY通道和Mighlos的不同相互作用引起可测量的电导差异，从而为不同的残基带来了可解释的签名，以进行测序。 ONT是一家全球成功的公司，拥有大量的技术创新。因此，该合作伙伴关系将汇集SEC机械的专业知识，并结合测序知识和平台，为您的项目成功，培训以及学术和生物技术环境和团队合作的经验提供了理想的环境。