Abstract
Reconstituting membrane proteins in liposomes and determining their structure is a common method for determining membrane protein structures using single-particle cryo-electron microscopy(cryo-EM).However,the strong signal of li-posomes under cryo-EM imaging conditions often interferes with the structural determination of the embedded membrane proteins.Here,we propose a liposome signal subtraction method based on single-particle two-dimensional(2D)classifica-tion average images,aimed at enhancing the reconstruction resolution of membrane proteins.We analyzed the signal distri-bution characteristics of liposomes and proteins within the 2D classification average images of protein-liposome complexes in the frequency domain.Based on this analysis,we designed a method to subtract the liposome signals from the original particle images.After the subtraction,the accuracy of single-particle three-dimensional(3D)alignment was improved,enhancing the resolution of the final 3D reconstruction.We demonstrated this method using a PIEZO 1-proteoliposome dataset by improving the resolution of the PIEZO 1 protein.
基金项目
National Natural Science Foundation of China(32241023)
National Natural Science Foundation of China(92254306)
Fund from the Tsinghua-Peking Joint Center for Life sciences()
Beijing Frontier Research Center for Biological Structure()
NETL
NSTL
万方数据