Biology subjects

Lemire, S.

Publications and source records attributed to Lemire, S..

5 recordsLinked to original sources

bioRxiv2026

Insights into Genome Ejection by a Therapeutic phiKMV-like Bacteriophage

Ar-KM is a phiKMV-like therapeutic bacteriophage used in clinical candidate phage therapy cocktails to treat lung infections caused by Pseudomonas aeruginosa. Here, we present an integrative structural atlas of Ar-KM proteins using cryo-EM, proteomics, and bioinformatics. From a single purified Ar-KM preparation, we identified three distinct populations: mature DNA-filled virions, open-nozzle particles with ejection proteins extending from the tail, and closed-nozzle empty particles. Near-atomic-resolution reconstructions of all three states enabled us to build atomic models for eleven structural proteins. The mature virion revealed the pre-ejection conformation of three ejection proteins, gp41, gp42, and gp43, homologous to coliphage T7s gp14, gp15, and gp16, respectively. Unlike T7, peptidoglycan hydrolase activity associated with the ejectosome resides in the gp15-like periplasmic tunnel protein gp42, whereas in T7 the lysozyme-like domain is located at the N-terminus of gp16, underscoring the structural plasticity and evolutionary mosaicity of ejection proteins. We further identified a short -helical factor, gp34, present in eight copies at the mismatched interface between the portal barrel and gp41. Gp34 forms a cage within the nozzle, acting as a molecular wedge that stabilizes the open conformation and permits gp41 to assemble into a hexameric channel during ejection. Evolutionarily, gp34 appears to be an ortholog of the essential gene gp7.3 in phage T7 and is conserved across sequenced phiKMV-like phages. We propose that this protein functions as an ejection protein assembly factor, stabilizing the open nozzle during infection and allowing the coordinated exit of ejection proteins and their assembly into a DNA-ejectosome. HighlightsO_LIStructural polymorphism of phage Ar-KM defines open and closed nozzle states C_LIO_LIEjection proteins gp43, gp42, and gp41 assemble in a 4:8:8 stoichiometry C_LIO_LILysozyme domain shows modular positioning within the ejectosome C_LIO_LIPortal protein acts as the primary barrier to genome leakage C_LIO_LIWedge protein gp34 (T7 gp7.3 ortholog) bridges portal and gp41 C_LI

biophysics↗

bioRxiv2025

Structural atlas of Pakpunavirus P7-1 reveals determinants of virion stability and genome ejection

Bacteriophages of the Pakpunavirus genus exhibit broad host range and potent bacteriolytic activity, making them promising candidates for clinical use. Here, we present a structural atlas of the therapeutic phage Pakpunavirus P7-1, a component of a phage cocktail targeting Pseudomonas aeruginosa that has undergone Phase 1/2 clinical trials. We determined the near-atomic structure of the extended virion and obtained a medium-resolution reconstruction of the contracted tail. Atomic models were built for 20 structural proteins comprising the icosahedral capsid, neck, contractile tail, and baseplate. We identified six upward-pointing Short Tail Fibers that stabilize the extended sheath and six highly flexible Long Tail Fibers likely involved in host recognition. Ordered fragments of the Tape Measure Protein revealed six copies inside the tail tube, forming a 3-helix cork at the tail tip. Sheath contraction repositions the baseplate, projecting all twelve tail fibers outward, yet contraction alone is insufficient to trigger genome ejection.

biochemistry↗

bioRxiv2025

High-resolution cryo-EM analysis of the therapeutic Pseudomonas phage Pa223

Cryogenic electron microscopy (cryo-EM) analysis of bacteriophages is a valuable method for deciphering virus composition and conformational plasticity. In this study, we present a high-resolution structural atlas of the Pseudomonas virus Pa223, a phage from the Bruynoghevirus genus that has recently been used in clinical cocktails for treating cystic fibrosis and non-cystic fibrosis bronchiectasis, as well as for compassionate care. By combining bioinformatics, proteomics, cryo-EM single particle analysis, and localized reconstruction, we annotated and built atomic models for eight structural polypeptide chains that form the icosahedral capsid and noncontractile tail. We discovered that the Pa223 capsid is decorated by a spike protein that features a unique triple-{beta} helix fold with no structural homologs in the database. The Pa223 tail features six trimeric tail fibers extending upwards, similar to, but shorter than, those found in phage T7. Unlike T7, the Pa223 tail is extended by two head-to-tail adaptors and sealed by a trimeric tail needle, similar to P22-like phages. We identified a protein bound around the outer perimeter of the portal protein, positioned similarly to the ejection protein gp72, which was identified in the Pseudomonas phage DEV, a Litunavirus phage and member of the reclassified Schitoviridae family. This structural hint led us to identify the Pa223 ejection proteins gp53, gp54, and gp56, which bioinformatically resemble those of T7-like phages more closely than Schitoviridae. Thus, phage Pa223 contains diverse structural elements found in P22-like, T7-like, and Litunavirus phages, providing a framework for understanding the diversification and evolution of ejection proteins in Bruynogheviruses. HighlightsO_LIThe high-resolution structure of Bruynoghevirus Pa223 reveals hybrid structural features that are shared among P22-like, T7-like, and Litunavirus phages. C_LIO_LIThe Pa223 capsid is decorated with a trimeric spike asymmetrically bound at the icosahedral 3-fold axes. C_LIO_LIThe Pa223 tail features two quasi-equivalent conformations of the head-to-tail adaptor protein arranged into two coaxial rings. C_LIO_LIIdentification of the ejection protein gp54 through structural similarity to gp72 from the Litunavirus DEV. C_LIO_LIBioinformatic mapping of the Pa223 ejection proteins gp53 and gp56 validated through mass spectrometry analysis of infectious virions. C_LI

biochemistry↗

bioRxiv2025

Exercise-Induced Myostimulin Enhances Muscle Function in Health and Disease

Musculoskeletal diseases are a leading contributor to years lived with disability worldwide1,2. While exercise offers significant benefits for people with these conditions, many individuals do not engage in adequate physical activity3. Consequently, there is growing interest in pharmacological interventions that can emulate essential health-promoting effects of exercise4,5. By integrating transcriptomics data of exercised skeletal muscle, we identified C1orf54/C1ORF54 as a novel exercise-responsive gene in mice and humans. We demonstrate that removal of the first sixteen N-terminal amino acids of C1ORF54 gives rise to a previously uncharacterized protein that stimulates the proliferation of muscle precursor cells and which we named myostimulin. Intriguingly, repeated intermittent treatment of mice with recombinant myostimulin boosts maximal isometric strength in mice within a week. Moreover, we have engineered a variant with improved biophysical properties, increased biological activity in vitro and enhanced efficacy in vivo. This variant even accelerates the recovery of muscle strength from axonotmesis, a condition associated with pronounced muscle weakness. Our data ascribe to myostimulin a role for enhancing the regenerative capacity of skeletal muscle and mediating functional adaptations characteristic of sustained resistance training. Therefore, myostimulin could be an innovative, fast acting therapeutic for certain human musculoskeletal diseases, injuries and other disorders that improve with exercise.

physiology↗

bioRxiv2019

Engineering Phage Host-Range and Suppressing Bacterial Resistance Through Phage Tail Fiber Mutagenesis

The rapid emergence of antibiotic-resistant infections is prompting increased interest in phage-based antimicrobials. However, acquisition of resistance by bacteria is a major issue in the successful development of phage therapies. Through natural evolution and structural modeling, we identified host-range determining regions (HRDR) in the T3 phage tail fiber protein and developed a high-throughput strategy to genetically engineer these regions through site-directed mutagenesis. Inspired by antibody specificity engineering, this approach generates deep functional diversity (>107 different members), while minimizing disruptions to the overall protein structure, resulting in synthetic \"phagebodies\". We showed that mutating HRDRs yields phagebodies with altered host-ranges. Select phagebodies enable long-term suppression of bacterial growth by preventing the appearance of resistance in vitro and are functional in vivo using a mouse skin infection model. We anticipate this approach may facilitate the creation of next-generation antimicrobials that slow resistance development and could be extended to other viral scaffolds for a broad range of applications.\n\nHighlightsO_LIVastly diverse phagebody libraries containing 107 different members were created.\nC_LIO_LIStructure-informed engineering of viral tail fibers efficiently generated host-range alterations.\nC_LIO_LIPhagebodies prevented the development of bacterial resistance across long timescales in vitro and are functional in vivo.\nC_LI

synthetic biology↗