Chemical Protein Glycosylation

描述（由申请人提供）：本资助申请旨在研究调节基本蛋白质生物物理特性的生物化学和生物医学意义（例如，结构、动力学、稳定性和功能）。主要假设是基于以下事实：化学糖基化有效地降低蛋白质结构动力学，从而改变其他相关蛋白质性质（例如，酶动力学和热力学稳定性）。这项新技术的发展将允许探索与蛋白质结构动力学相关的许多基本科学问题。由于蛋白质动力学可以通过增加糖基化水平逐渐改变，我们将能够首次确定蛋白质结构动力学如何影响这些其他性质。多种生物物理特性将被确定为糖基化的函数，并且它们的变化具有统计学相关性。此外，分子建模和动力学模拟技术将被用来探索聚糖实现这种效果的分子机制。因此，这些实验将提供有关动态对蛋白质中所谓的结构-功能和结构-稳定性关系的影响的基本见解。此外，我们将研究生物医学相关应用中通过化学糖基化稳定蛋白质的基本方面，即液态和固态制剂、缓释装置以及与生物和纳米材料的相互作用（SA 2 -4）。功能稳定性参数（例如，失活，聚集体形成）将与先天生物物理性质（例如，结构动力学，热力学稳定性）和化学糖基化变量（例如，聚糖的大小和化学性质、糖基化水平）。实验旨在建立化学糖基化增强功能稳定性的机制。因此，这项工作的直接生物医学相关性在于进一步理解蛋白质热力学、动力学和胶体稳定性在治疗应用中可以增加的机制。这项研究的长期目标包括进一步发展化学糖基化，用于研究蛋白质结构动力学和预防蛋白质不稳定性（例如，失活、聚集）。项目相关性。蛋白质疗法的临床适用性关键取决于保持其功能和结构的完整性。通过固体和液体制剂中的化学糖基化以及暴露于界面（例如，塑料管、医疗设备的表面），对于确定治疗的最大效率和防止不利的副作用（例如，由施用聚集蛋白引起的免疫反应）。我们的研究将为增强蛋白质的热力学和动力学稳定性提供新的策略。

DESCRIPTION (provided by applicant): This grant application seeks to study the biochemical and biomedical implications of modulating fundamental protein biophysical properties (e.g., structure, dynamics, stability, and function) through chemical glycosylation. The main hypothesis is based on the fact that chemical glycosylation effectively reduces protein structural dynamics thereby altering other relevant protein properties (e.g., enzyme kinetics and thermodynamic stability). The development of this novel technology will allow the exploration of a multitude of fundamental scientific questions related to protein structural dynamics. Since the protein dynamics can be shifted gradually by increasing the glycosylation level, we will be able to determine for the first time how protein structural dynamics influences these other properties. Multiple biophysical properties will be determined as a function of glycosylation and their changes statistically correlated. Additionally, molecular modeling and dynamics simulation techniques will be employed to explore the molecular mechanisms by which glycans achieve such effects. These experiments will thus provide fundamental insights regarding the influence of dynamics on the so-called structure-function and structure-stability relationships in proteins. Furthermore, we will study fundamental aspects regarding the stabilization of proteins by chemical glycosylation within biomedically relevant applications, namely, liquid- and solid-state formulations, sustained release devices, and interactions with bio- and nano-materials (SA2-4). Functional stability parameters (e.g., inactivation, aggregate formation) will be correlated with the innate biophysical properties (e.g., structural dynamics, thermodynamic stability) of the glycoconjugates and with the chemical glycosylation variables (e.g., glycan's size and chemical nature, glycosylation level). Experiments are designed to establish the mechanisms of enhanced functional stability by chemical glycosylation. The direct biomedical relevance of the work thus consists in furthering the understanding of the mechanisms by which protein thermodynamic, kinetic, and colloidal stabilities can be increased within therapeutic applications. The long-term goals of this research consist on furthering the development of chemical glycosylation for the study of protein structural dynamics and for the prevention of protein instabilities (e.g., inactivation, aggregation) during storage and delivery of protein therapeutics. Project Relevance. The clinical applicability of protein therapeutics critically depends on preserving their functional and structural intactness. Increasing protein stability by chemical glycosylation in solid and liquid formulations and upon exposure to interfaces (e.g., plastic tubing, surfaces of medical equipment), is essential to ascertain maximum efficiency of the treatment and preventing adverse side effects (e.g., immune reactions caused by administration of aggregated protein). Our research will provide new strategies to enhance protein thermodynamic and kinetic stability.