Localization of the inner body in jumbo phage phiK601 by Cryo-EM and high-dose imaging

Andrey V. Moiseenko; Ishika Gupta; Grigorii Mitrov; Mikhail Egorochkin; Ruqaiyah Khan; Virajith Boddapati; Sayani Das; Sandip Kaledhonkar; Kiran Kondabagil; Olga S. Sokolova; Andrey V. Moiseenko; Ishika Gupta; Grigorii Mitrov; Mikhail Egorochkin; Ruqaiyah Khan; Virajith Boddapati; Sayani Das; Sandip Kaledhonkar; Kiran Kondabagil; Olga S. Sokolova

doi:10.3934/biophy.2026005

AIMS Biophysics

2026, Volume 13, Issue 1: 80-93. doi: 10.3934/biophy.2026005

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Localization of the inner body in jumbo phage phiK601 by Cryo-EM and high-dose imaging

1.
Faculty of Biology, Lomonosov Moscow State University, Moscow, 119234, Russia
2.
Department of Biosciences and Bioengineering, Indian Institute of Technology Bombay, Mumbai, 400076, India
3.
Koita Centre for Digital Health (KCDH), Indian Institute of Technology Bombay, Mumbai, 400076, India
4.
Faculty of Biology, Shenzhen MSU-BIT University, Shenzhen, 518172, China
5.
Key Laboratory of Forest Plant Ecology, Ministry of Education, Northeast Forestry University, Harbin, 150040, China

Received: 12 November 2025 Revised: 12 February 2026 Accepted: 24 February 2026 Published: 16 March 2026

Jumbo bacteriophages possess exceptionally large capsids accommodating genomes that encode additional proteins, which support their infection and replication. A distinctive structural element, known as the inner body, has been observed in a number of phiKZ-like phage particles and its proteins are believed to play an essential role in phage genome organization and ejection. However, the precise localization and three-dimensional structure of the inner body have remained elusive. Here, we applied the high-dose cryo-electron microscopy (“bubblegram”) approach to localize the inner body within the capsid of phiKZ-like jumbo phage phiK601. The inner body was resolved as a cylindrical structure approximately 22 nm in diameter, tilted by ~ 20° relative to the tail axis and positioned asymmetrically, likely contacting the portal vertex and the opposing capsid edge. It is surrounded by 17 concentric layers of packaged DNA and exhibits positional flexibility within the capsid.
- jumbo phage,
- cryo-electron microscopy,
- inner body,
- bubblegrams,
- phiK601
Citation: Andrey V. Moiseenko, Ishika Gupta, Grigorii Mitrov, Mikhail Egorochkin, Ruqaiyah Khan, Virajith Boddapati, Sayani Das, Sandip Kaledhonkar, Kiran Kondabagil, Olga S. Sokolova. Localization of the inner body in jumbo phage phiK601 by Cryo-EM and high-dose imaging[J]. AIMS Biophysics, 2026, 13(1): 80-93. doi: 10.3934/biophy.2026005

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Abstract

Jumbo bacteriophages possess exceptionally large capsids accommodating genomes that encode additional proteins, which support their infection and replication. A distinctive structural element, known as the inner body, has been observed in a number of phiKZ-like phage particles and its proteins are believed to play an essential role in phage genome organization and ejection. However, the precise localization and three-dimensional structure of the inner body have remained elusive. Here, we applied the high-dose cryo-electron microscopy (“bubblegram”) approach to localize the inner body within the capsid of phiKZ-like jumbo phage phiK601. The inner body was resolved as a cylindrical structure approximately 22 nm in diameter, tilted by ~ 20° relative to the tail axis and positioned asymmetrically, likely contacting the portal vertex and the opposing capsid edge. It is surrounded by 17 concentric layers of packaged DNA and exhibits positional flexibility within the capsid.

Acknowledgments

Authors thank Mrs Lisa Trifonova for proof-reading the manuscript. The cryo-EM reconstruction was supported by RSF (24-44-02003 to O.S.S.) and the phage isolation and purification–by Department of Science and Technology, India (DST/INT/RUS/RSF/P-79/2023 to K.K.). K.K. lab was also supported partly by funding from the Indian Council of Medical Research, ICMR (IIRPSG-2024-01-02611), and the Council of Scientific and Industrial Research (CSIR, 37/1752/23/EMR-II). S.K. lab acknowledges support from Science and Engineering Research Board, Government of India (IPA/2020/000413). We also thank the ANRF and IoE-supported national cryo-EM facility at IIT Bombay.), V.B. also acknowledges the fellowship from the Council of Scientific & Industrial Research (CSIR), India (Grant No: 09/0087(13984)/2022-EMR-I).

Conflict of interest

The authors declare no conflict of interest.

Author contributions

A.V.M. performed cryo-EM data processing and drafted the manuscript with input from all authors, M.E. and G.M. performed phage capsid reconstruction, I.G., R.K., and S.D. performed the phage purification, genomics, and virology studies, G.M. performed partial annotation of head and inner body proteins, S.K. supervised the cryo-electron microscopy data collection, V.B. performed cryo-electron microscopy data collection, K.K. and O.S.S. designed and supervised the study.

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