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[前沿资讯 ] 学者开发出基于氟磺酸的可富集化学交联剂 进入全文
科学网
近日，中国科学院广州生物医药与健康研究院研究员唐士兵与浙江大学研究员杨兵、北京航空航天大学副教授刘超团队合作，在国家自然科学基金、国家重点研发计划等项目的支持下，开发出一种新型氟磺酸类可富集化学交联的非天然氨基酸并在活细胞中研究蛋白质直接相互作用。相关成果发表于《自然-通讯》。 蛋白质-蛋白质相互作用鉴定是蛋白质功能研究的重要步骤，在活细胞中原位鉴定直接相互作用蛋白对于生物医药研究具有重要意义。基于邻近触发反应的化学交联非天然氨基酸可以捕获弱作用力和瞬时蛋白质相互作用，已被开发用于原位鉴定活细胞中的蛋白质相互作用。基于质谱的交联肽段解析能够提高互作蛋白鉴定的特异性、确定蛋白相互作用界面。然而，由于蛋白质样品和质谱数据解析过程的复杂性，对蛋白质化学交联后的交联产物进行高通量鉴定具有挑战性。在蛋白质样品进行质谱分析前，对交联肽段进行富集是提高鉴定效率的有效策略。
[前沿资讯 ] 中科院天津工业生物所开发面向CRISPR技术的基因组编辑自动化设计在线工具AutoESDcas 进入全文
中科院天津工业生物所
近日，中国科学院天津工业生物技术研究所生物设计中心平台实验室开发了面向CRISPR技术的基因组编辑自动化设计在线工具AutoESDcas（https://autoesdcas.biodesign.ac.cn）。该工具面向CRISPR介导的同源重组技术和Golden Gate组装技术，通过搭建自动化编辑序列设计流程，实现了基因组编辑的自动化和高通量设计；通过整合多种基因组编辑任务的设计流程，实现了对多种基因组编辑实验场景的支持；通过添加同源臂脱靶风险评估和基于脱靶分析的引物优化功能，提高了设计结果的可靠性。该工具能够自动化完成针对不同实验场景的基因组编辑实验流程所需的全套编辑序列设计。针对多个物种的数百个设计任务可在一小时内完成，尤其适用于面向高通量基因编辑平台的大规模基因组编辑设计。
[前沿资讯 ] Reversal of the histidine kinase activity by a small linker helix 进入全文
Eurekalert
Scientists have revealed new information on how red light-regulated histidine kinases function. According to the study, altering of the length of a specific ‘linker helix’ region of the protein can even reverse their enzymatic activity. The study is published in Nature Communications. An international collaboration between the groups of Dr. Heikki Takala from the University of Jyväskylä and Prof. Andreas Möglich from the University of Bayreuth have revealed key details on histidine kinase signaling. By applying modifications in their pREDusk tool, they showed how the delicate balance between kinase and phosphatase activities is fine-tuned in histidine kinase receptors. Importantly, they also revealed that certain deletions in a so-called ‘linker helix’ of the phytochrome component reverses their enzymatic activity.
[前沿资讯 ] P450酶催化糖肽分子内苯酚偶联反应机制获揭示 进入全文
科学网
近日，中国科学院南海海洋研究所研究员张长生团队和厦门大学教授王斌举团队合作，在细胞色素P450酶催化糖肽分子内苯酚偶联反应的机制研究方面取得新进展。这为发现和开发更多新颖的双环肽类化合物奠定了研究基础，有望为抗生素研发提供更多物质基础。相关成果发表于《美国化学会志》。 研究团队从放线菌Amycolatopsis cihanbeyliensis DSM 45679中发现了C−O连接的新型双环糖肽类化合物cihanmycins (CHMs)。CHM A (1)的结构通过X-ray单晶衍射确定为双环糖肽，包含肉桂酰基团和两个稀有的D-阿拉伯呋喃糖。研究团队基于生物信息学分析和异源表达实验，在DSM 45679的基因组中定位了CHM的基因簇，并通过体内基因敲除和体外生化实验证实了3个P450酶的功能：Cih26负责肉桂酰基团的环氧化和羟化，Cih32负责三个氨基酸的羟化，Cih33及其同源蛋白DmlH和EpcH催化分子内C−O苯酚偶联反应形成CHM的双环骨架。
[学术文献 ] Perspective on Agricultural Industrialization: Modification Strategies for Enhancing the Catalytic Capacity of Keratinase 进入全文
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
Keratinases is a special hydrolytic enzyme produced by microorganisms, which has the ability to catalyze the degradation of keratin. Currently, keratinases show great potential for application in many agricultural and industrial fields, such as biofermented feed, leather tanning, hair removal, and fertilizer production. However, these potentials have not yet been fully unleashed on an industrial scale. This paper reviews the sources, properties, and catalytic mechanisms of keratinases. Strategies for the molecular modification of keratinases are summarized and discussed in terms of improving the substrate specificity, thermostability, and pH tolerance of keratinases. The modification strategies are also enriched by the introduction of immobilized enzymes and directed evolution. In addition, the selection of modification strategies when facing specific industrial applications is discussed and prospects are provided. We believe that this review serves as a reference for the future quest to extend the application of keratinases from the laboratory to industry.
[学术文献 ] High-throughput system for the thermostability analysis of proteins 进入全文
Protein Science
Thermal stability of proteins is a primary metric for evaluating their physical properties. Although researchers attempted to predict it using machine learning frameworks, their performance has been dependent on the quality and quantity of published data. This is due to the technical limitation that thermodynamic characterization of protein denaturation by fluorescence or calorimetry in a high-throughput manner has been challenging. Obtaining a melting curve that derives solely from the target protein requires laborious purification, making it far from practical to prepare a hundred or more samples in a single workflow. Here, we aimed to overcome this throughput limitation by leveraging the high protein secretion efficacy of Brevibacillus and consecutive treatment with plate-scale purification methodologies. By handling the entire process of expression, purification, and analysis on a per-plate basis, we enabled the direct observation of protein denaturation in 384 samples within 4 days. To demonstrate a practical application of the system, we conducted a comprehensive analysis of 186 single mutants of a single-chain variable fragment of nivolumab, harvesting the melting temperature (Tm) ranging from −9.3 up to +10.8°C compared to the wild-type sequence. Our findings will allow for data-driven stabilization in protein design and streamlining the rational approaches.
