PDF(Pubmed)
Abstract：
Understanding protein structure and kinetics under physiological conditions is crucial for elucidating complex biological processes. While time-resolved (TR) techniques have advanced to track molecular actions, their practical application in biological reactions is often confined to reversible photoreactions within limited experimental parameters due to inefficient sample utilization and inflexibility of experimental setups. Here, we introduce serial X-ray liquidography (SXL), a technique that combines time-resolved X-ray liquidography with a fixed target of serially arranged microchambers. SXL breaks through the previously mentioned barriers, enabling microgram-scale TR studies of both irreversible and reversible reactions of even a non-photoactive protein. We demonstrate its versatility in studying a wide range of biological reactions, highlighting its potential as a flexible and multi-dimensional assay framework for kinetic and structural characterization. Leveraging X-ray free-electron lasers and micro-focused X-ray pulses promises further enhancements in both temporal resolution and minimizing sample quantity. SXL offers unprecedented insights into the structural and kinetic landscapes of molecular actions, paving the way for a deeper understanding of complex biological processes.
摘要：
了解生理条件下的蛋白质结构和动力学对于阐明复杂的生物过程至关重要。虽然时间分辨（TR）技术已经发展到跟踪分子作用，它们在生物反应中的实际应用通常局限于在有限的实验参数内的可逆光反应，这是由于样品利用率低和实验设置不灵活。这里,我们介绍了串行X射线液相成像(SXL)，一种将时间分辨X射线液相成像与串联排列的微室的固定目标相结合的技术。SXL突破了前面提到的障碍，即使是非光敏蛋白的不可逆反应和可逆反应，也可以进行微克规模的TR研究。我们证明了它在研究广泛的生物反应方面的多功能性，强调其作为动力学和结构表征的灵活和多维测定框架的潜力。利用X射线自由电子激光器和微聚焦X射线脉冲有望进一步增强时间分辨率和最大限度地减少样本量。SXL为分子作用的结构和动力学景观提供了前所未有的见解，为更深入地了解复杂的生物过程铺平道路。
