文献类型：期刊文献

英文题名：Optimized Tandem Affinity Purification Strategy Enables High-Yield Isolation and Functional Characterization of Native COMPASS in Saccharomyces cerevisiae

作者：Li, Ya[1];Quan, Shu[2,3];Zheng, Yongxin[1,2,3]

机构：[1]East China Univ Sci & Technol, Shanghai Frontiers Sci Ctr Optogenet Tech Cell Met, Sch Biotechnol, Shanghai 200237, Peoples R China;[2]Shanghai Jiao Tong Univ, Sch Life Sci & Biotechnol, State Key Lab Microbial Metab, Shanghai 200240, Peoples R China;[3]Shanghai Jiao Tong Univ, Zhangjiang Inst Adv Study, Shanghai 201203, Peoples R China

年份：2025

卷号：15

期号：6

外文期刊名：CATALYSTS

收录：;EI(收录号：20252618672265);WOS:【SCI-EXPANDED(收录号:WOS:001515481600001)】；

基金：This research was funded by the National Natural Science Foundation of China (NSFC), Grant Numbers: 32301059 and 32222049. Shanghai Municipal Science and Technology Major Project; The Shanghai Frontier Science Center of Optogenetic Techniques for Cell Metabolism, Shanghai Municipal Education Commission, Grant Number: 2021 Sci & Tech 03 28.

语种：英文

外文关键词：COMPASS; Set1; methyltransferase; tandem affinity purification; budding yeast

摘要：Histone 3 lysine 4 methylation (H3K4me) is an evolutionarily conserved epigenetic marker associated with transcriptional activation, playing a crucial role in growth and development. In yeast, all forms of H3K4 methylation are catalyzed by the COMPASS complex. However, purifying endogenous COMPASS remains challenging due to its low abundance, compositional complexity, and structural instability, resulting in low yield, poor purity, and heterogeneity in isolated complexes. These technical limitations have impeded the structural elucidation of the intact COMPASS complex and contributed to inconsistencies in reported in vitro enzymatic activity, thereby limiting a comprehensive understanding of its functions. Here, we present an optimized tandem affinity purification strategy that enables the high-yield isolation of native COMPASS from Saccharomyces cerevisiae with >99% purity and intact subunit composition, as validated by biochemical analyses. Using recombinant nucleosomes as substrates, we systematically characterized its catalytic properties and found that endogenously purified COMPASS exhibited strict dependence on H2B ubiquitination for catalyzing H3K4 methylation. This work establishes an efficient purification strategy for future structural and functional studies of COMPASS and provides critical insights into its catalytic properties.