A hydrogen–deuterium exchange mass spectrometry-based protocol for protein–small molecule interaction analysis  

作　　者：Qian Meng Yuan-Li Song Chen Zhou Han He Naixia Zhang Hu Zhou 

机构地区：[1]Analytical Research Center for Organic and Biological Molecules,State Key Laboratory of Drug Research,State Key Laboratory of Chemical Biology,Shanghai Institute of Materia Medica,Chinese Academy of Sciences,Shanghai 201203,China [2]University of Chinese Academy of Sciences,Beijing 100049,China [3]Department of Pharmacology,School of Pharmacy,Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China [4]School of Pharmaceutical Science and Technology,Hangzhou Institute for Advanced Study,University of Chinese Academy of Sciences,Hangzhou 310024,China

出　　处：《Biophysics Reports》2023年第2期99-111,共13页生物物理学报（英文版）

基　　金：supported by the National Key Research and Development Program(2022YFA1302900);Shanghai Young Excellent Academic Leader Program(20XD1424900);Shanghai Municipal Science and Technology Major Project;the National Natural Science Foundation of China(32171220,22107111,21977105);the Youth Innovation Promotion Association CAS(2022284)。

摘　　要：Protein–small molecule interaction is vital in regulating protein functions and controlling various cellular processes.Hydrogen deuterium exchange mass spectrometry(HDX-MS)is a powerful methodology to study protein–small molecule interactions,however,to accurately probe the conformational dynamics of the protein upon small molecule binding,the HDX-MS experimental conditions should be carefully controlled and optimized.Here,we present the detailed continuous-labeling,bottom-up HDX-MS protocol for studying protein–small molecule interactions.We took a side-by-side HDX kinetics comparison of the Hsp90N protein with or without the treatment of small molecules(i.e.,Radicicol,Geldanamycin)for displaying conformational changes induced by molecular interactions between Hsp90N and small molecules.Our sensitive and robust experimental protocol can facilitate the novice to quickly carry out the structural characterization of protein–small molecule interactions.

关 键 词：Hydrogen–deuterium exchange mass spectrometry(HDX-MS) Protein dynamics Protein–small molecule interactions Hsp90N 

分 类 号：Q51[生物学—生物化学]