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In this video (Part 4 of 6), we investigate Encapsulated Ferritin (EncFer, EMPIAR 10716: https://www.ebi.ac.uk/empiar/EMPIAR-1.... Each particle in this cryo-EM dataset has two parts: an icosahedral encapsulin and four D5 symmetric EncFer proteins. The EncFer are contained inside the encapsulin protein, meaning there is a symmetry mismatch between the outer shell and the inner parts. The majority of the case study focuses on improving the resolution of the internal EncFer molecules. Interestingly, although four EncFer molecules appear tetrahedral, their internal D5 symmetry means that the overall arrangement has no point-group symmetry. This requires the use of two new workflows in CryoSPARC, which we call Group Re-alignment and Custom Symmetry Expansion. A written companion to this video is available in the CryoSPARC Guide. See: Case Study: End-to-end processing of encapsulated ferritin: https://guide.cryosparc.com/processin... Chapters: 00:00 Introduction 02:21 Symmetry in Cryo-EM 07:58 EncFer Background 10:06 Symmetry Mismatch 20:59 Encapsulin map 25:15 Particle Subtraction and Refinement of EncFer 33:27 Q&A 35:09 Volume Alignment Methods 41:23 Group Re-alignment 45:13 Custom Symmetry Expansion 1:01:45 Q&A 1:15:37 Local Refinement and Final Map 1:30:59 Final Q&A This video is the fourth in a series of six recordings covering single particle cryo-EM data processing in CryoSPARC from the 2024 Single-Particle Cryo-EM Image Processing Workshop held by the Stanford-SLAC CryoEM Center (S2C2) in 2024. The original recordings are courtesy of S2C2. Captions were automatically generated by Zoom and may contain errors. Workshop: https://s2c2.slac.stanford.edu/traini... S2C2: https://s2c2.slac.stanford.edu/traini... More tutorials and videos are available in the CryoSPARC Guide: https://guide.cryosparc.com/