REL 2023-11-27 2024-03-26 2024-03-26 0000-0001-6656-6320 Yong Zi Tan National University of Singapore 14 Science Drive 4 Singapore Singapore Singapore 117543 0000-0001-6656-6320 Yong Zi Tan National University of Singapore 14 Science Drive 4 Singapore Singapore Singapore 117543 0000-0002-8124-7472 Dmitry Lyumkis Salk Institute 10010 N. Torrey Pines Rd. San Diego California United States 92037 Aiyer S Baldwin PR Tan SM Shan Z Oh J Mehrani A Bowman ME Louie G Passos DO Đorđević-Marquardt S Mietzsch M Hull JA Hoshika S Barad BA Grotjahn DA McKenna R Agbandje-McKenna M Benner SA Noel JAP Wang D Tan YZ Lyumkis D National Institutes of Health (NIH) U54 AI170855 United States National Institutes of Health (NIH) U01 AI136680 United States National Institutes of Health (NIH) GM148476 United States National Institutes of Health (NIH) GM082946 United States National Science Foundation (NSF, United States) MCB 2048095 United States Other private Hearst Foundations Developmental Chair United States Other private Margaret T. Morris Foundation United States National Research Foundation (NRF, Singapore) NRF Fellowship A-8001346-00-00 Singapore Other government National University of Singapore (PYP Fellowship A-0008405-00-00, A-0008405-01-00) Singapore Ministry of Education (MoE, Singapore) MOE AcRF Tier 1 A-8000037-00-00 Singapore Ministry of Education (MoE, Singapore) MOE AcRF Tier 2 A-6100427-01-00 Singapore Other government Agency for Science, Technology and Research, Singapore Singapore Damon Runyon Cancer Research Foundation Nadia’s Gift Foundation Innovator Award DRR-65-21 United States American Cancer Society PF-21-075-01-CCB United States Other government Cancer Center Support Grant P30 CA01495t United States National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS) GM103310 United States Simons Foundation SF349247 United States Other government NYSTAR United States Other government S10 OD032467 United States National Science Foundation (NSF, United States) MCB-1939086 United States 487.4 10.6019/EMPIAR-11797 EMDB molecule EMD-41695 Aiyer S Baldwin PR Tan SM Shan Z Oh J Mehrani A Bowman ME Louie G Passos DO Đorđević-Marquardt S Mietzsch M Hull JA Hoshika S Barad BA Grotjahn DA McKenna R Agbandje-McKenna M Benner SA Noel JAP Wang D Tan YZ Lyumkis D bioRxiv 10.1101/2023.07.14.548955 Aiyer S Baldwin PR Tan SM Shan Z Oh J Mehrani A Bowman ME Louie G Passos DO Đorđević-Marquardt S Mietzsch M Hull JA Hoshika S Barad BA Grotjahn DA McKenna R Agbandje-McKenna M Benner SA Noel JAP Wang D Tan YZ Lyumkis D Nature communications Nat Commun 1 15 2024 English 10.1038/s41467-023-44555-7 38195598 Unaligned multiframe movies of RNA polymerase collected at various tilts /data/movies micrographs - multiframe TIFF TIFF 1146 51 1 50 UNSIGNED BYTE 4092 0.83 5760 0.83

Movies collected using Titan Krios and K3 quantum with energy filter. Tilt angles of 0, 10, 20, 30, 40, 50 and 60 degrees were collected put into their respective folders. 50 frames were used, pixel size is 0.830 Å/pixel, total dose of 34.2 e-/Å2 (0 degrees) and 38.8 e-/Å2 (60 degrees). Gain reference is 22nov04c_04153642_01_4092x5760_norm_0.mrc.

data/micrographs/T*0/*mrc data/stack/RNAP-T*0-Polished.star data/stack Aligned and dose-weighted micrographs of RNA polymerase collected at various tilts /data/micrographs micrographs - single frame MRC MRC 1146 1 32 BIT FLOAT 4092 0.83 5760 0.83

Aligned and dose weighted using UCSF MotionCor2. Tilt angles of 0, 10, 20, 30, 40, 50 and 60 degrees were collected put into their respective folders.

data/micrographs/T*0/*mrc data/stack/RNAP-T*0-Polished.star data/stack Polished and equalized particles of RNA polymerase collected at various tilts /data/stack picked particles - multiframe - processed MRCS MRCS 2 72000 1 72000 32 BIT FLOAT 312 0.83 312 0.83

Final particle stack after particle polishing using Relion. The particles are equalized so that there are same number of particles per tilt angle with similar defocus distribution.

data/micrographs/T*0/*mrc data/stack/RNAP-T*0-Polished.star data/stack