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ORIGINAL RESEARCH article

Front. Microbiol.

Sec. Phage Biology

Volume 16 - 2025 | doi: 10.3389/fmicb.2025.1550594

Characterization of Holins, The Membrane Proteins of Coliphage ASEC2201: A Genomewide In Silico Approach

Provisionally accepted
  • 1Amity Institute of Biotechnology, Amity University, Lucknow, Lucknow, India
  • 2Dr.Ram Manohar Lohia Institute Of Medical Sciences, Lucknow, Uttar Pradesh, India
  • 3Integral Institute of Agriculture Science and Technology, Integral University, Lucknow, Uttar Pradesh, India
  • 4Department of Bioengineering, Faculty of Engineering and Information Technology, Integral University, Lucknow, India
  • 5Faculty of Medicine, Alatoo International University, Bishkek, Kyrgyzstan

The final, formatted version of the article will be published soon.

Drug-resistant Escherichia coli poses a significant healthcare burden, driving the search for novel antimicrobials. Here, we report the isolation and whole-genome sequencing of ASEC2201, a novel coliphage derived from multidrug-resistant clinical E. coli strains. Genome annotation using Prokka identified three putative holin genes (PROKKA_03659, PROKKA_04292, and PROKKA_04422) belonging to the Phage_holin_2_1 superfamily. Upstream promoter prediction revealed active regulatory elements at positions 112, 177, and 186 for these genes, indicating robust transcriptional activity. Transmembrane topology analysis using DeepTMHMM confirmed the presence of two to three α-helical membrane-spanning domains in each holin, essential for pore formation. Homology modeling with SWISS-MODEL yielded high-confidence three-dimensional structures characterized by conserved membrane-anchoring motifs, as supported by QMEAN and GMQE quality scores. In silico identification of cell-penetrating peptide motifs within the holin sequences suggests potential for enhanced intracellular delivery in CPP-fusion therapeutic constructs. Overall, our in-depth analysis elucidates the structural and functional properties of ASEC2201 holins, underscoring their biotechnological significance as scaffolds for developing novel antimicrobial strategies against MDR E. coli. It gives us an understanding on how the holins, with their inherent membrane-disrupting functions, can be explored in detail for future use as lysis modules in programmable bacterial systems, while their identified CPP motifs offer additional potential for engineering targeted therapeutic delivery vehicles. This study also demonstrates the potential of integrative in silico approaches in developing a comprehensive foundation for future experimental validation for proteins with no prior functional annotation

Keywords: transmembrane domain, Phage-mediated lysis, ASEC2201, protein 3D structure, functional annotation, Holins, Bacteriophage, Bacteriophage-encoded enzymes

Received: 23 Dec 2024; Accepted: 04 Jun 2025.

Copyright: © 2025 Singh, Saeed, Padmesh, Singh, Sen, Siddiqui, Hussain and Beg. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

* Correspondence: Aditi Singh, Amity Institute of Biotechnology, Amity University, Lucknow, Lucknow, India

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