A strategy combining denoising and cryo-EM single particle analysis.

In cryogenic electron microscopy (cryo-EM) single particle analysis (SPA), high-resolution three-dimensional structures of biological macromolecules are determined by iteratively aligning and averaging a large number of two-dimensional projections of molecules. Since the correlation measures are sensitive to the signal-to-noise ratio, various parameter estimation steps in SPA will be disturbed by the high-intensity noise in cryo-EM. However, denoising algorithms tend to damage high frequencies and suppress mid- and high-frequency contrast of micrographs, which exactly the precise parameter estimation relies on, therefore, limiting their application in SPA. In this study, we suggest combining a cryo-EM image processing pipeline with denoising and maximizing the signal's contribution in various parameter estimation steps. To solve the inherent flaws of denoising algorithms, we design an algorithm named MScale to correct the amplitude distortion caused by denoising and propose a new orientation determination strategy to compensate for the high-frequency loss. In the experiments on several real datasets, the denoised particles are successfully applied in the class assignment estimation and orientation determination tasks, ultimately enhancing the quality of biomacromolecule reconstruction. The case study on classification indicates that our strategy not only improves the resolution of difficult classes (up to 5 Å) but also resolves an additional class. In the case study on orientation determination, our strategy improves the resolution of the final reconstructed density map by 0.34 Å compared with conventional strategy. The code is available at https://github.com/zhanghui186/Mscale.