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Determining protein structures in cellular lamella at pseudo-atomic resolution by GisSPA

Jing Cheng,Tong Liu,X. You,Fa Zhang,S. Sui,Xiaohua Wan,Xinzheng Zhang

Published 2023 in Nature Communications

ABSTRACT

High-resolution in situ protein structure can be solved by cryo-ET, which requires several days of data collection. Here Cheng et al. report GisSPA, a program that may enable determining sub-4 Å resolution structures on cellular lamellae within one day of data collection. Cryo-electron tomography is a major tool used to study the structure of protein complexes in situ. However, the throughput of tilt-series image data collection is still quite low. Here, we show that GisSPA, a GPU accelerated program, can translationally and rotationally localize the target protein complex in cellular lamellae, as prepared with a focused ion beam, using single cryo-electron microscopy images without tilt-series, and reconstruct the protein complex at near-atomic resolution. GisSPA allows high-throughput data collection without the acquisition of tilt-series images and reconstruction of the tomogram, which is essential for high-resolution reconstruction of asymmetric or low-symmetry protein complexes. We demonstrate the power of GisSPA with 3.4-Å and 3.9-Å resolutions of resolving phycobilisome and tetrameric photosystem II complex structures in cellular lamellae, respectively. In this work, we present GisSPA as a practical tool that facilitates high-resolution in situ protein structure determination.

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