Quantitative time-resolved analysis reveals intricate, differential regulation of standard- and immuno-proteasomes

Proteasomal protein degradation is a key determinant of protein half-life and hence of cellular processes ranging from basic metabolism to a host of immunological processes. Despite its importance the mechanisms regulating proteasome activity are only incompletely understood. Here we use an iterative and tightly integrated experimental and modelling approach to develop, explore and validate mechanistic models of proteasomal peptide-hydrolysis dynamics. The 20S proteasome is a dynamic enzyme and its activity varies over time because of interactions between substrates and products and the proteolytic and regulatory sites; the locations of these sites and the interactions between them are predicted by the model, and experimentally supported. The analysis suggests that the rate-limiting step of hydrolysis is the transport of the substrates into the proteasome. The transport efficiency varies between human standard- and immuno-proteasomes thereby impinging upon total degradation rate and sub...蛋白酶体（proteasome）介导的蛋白质降解是决定蛋白质半衰期的关键因素，进而调控从基础代谢到各类免疫过程在内的一系列细胞生命活动。尽管其生物学意义重大，但目前对调控蛋白酶体活性的分子机制仍尚未完全阐明。本研究采用迭代式且紧密整合的实验与建模方法，构建、探究并验证了蛋白酶体肽水解动力学的机制模型。20S蛋白酶体（20S proteasome）是一类动态酶类，其活性随时间发生动态变化，这源于底物、产物与蛋白酶解位点及调控位点之间的相互作用；本模型预测了上述位点的分布及其相互作用模式，并得到了实验验证。分析结果表明，肽水解过程的限速步骤为底物向蛋白酶体内部的转运过程。人类标准蛋白酶体与免疫蛋白酶体之间的转运效率存在差异，进而影响整体降解速率及后续。