Identification and cryoEM Structure Determination of Escherichia Phage YDC107 Tail Found in a Bacteria-Contaminated Buffer DOI Creative Commons
Matthew C. Jenkins, Tapati Dutta, Daija Bobe

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Dec. 11, 2024

Abstract Cryo-electron microscopy data analysis can yield multiple structures from a single heterogeneous dataset. Here, we show workflow used for the identification of contaminant cryoEM grid without prior knowledge protein sequence. We determined tail structure Escherichia phage YDC107 only several thousand particles. The combines high-resolution single-particle processing with de novo model determination using ML-based methods. Structural revealed that central part has C6 symmetry, however overall symmetry each segment is C3 due to dimerization flexible domain. Figure

Language: Английский

Surface Engineering of the Encapsulin Nanocompartment of Myxococcus xanthus for Cell-Targeted Protein Delivery DOI Creative Commons

Sac Nicté Gómez-Barrera,

Willy Ángel Delgado-Tapia,

Aquetzali Estefanía Hernández-Gutiérrez

et al.

ACS Omega, Journal Year: 2025, Volume and Issue: 10(7), P. 7142 - 7152

Published: Feb. 12, 2025

Encapsulin nanocompartments (ENCs), or simply encapsulins, are a novel type of protein nanocage found in bacteria and archaea. The complete encapsulin systems include cargoes involved specific metabolic tasks. Cargoes selectively encapsulated due to the presence cargo-loading peptide (CLP). However, heterologous proteins fused CLP have also been successfully encapsulated, making encapsulins very promising system for protein-carrying delivery. Nevertheless, precise cell tissue delivery, require addition tagging peptides proteins. In this study, external surface Myxococcus xanthus ENC (MxENC) was analyzed modified carry bioorthogonal conjugation (SpyTag) further decorate MxENCs with any targeting previously SpyTag orthogonal pair, SpyCatcher protein. structural analysis MxENC led selection loop 155–159 C-terminus shell (EncA) genetic fusion peptide. engineered EncA forms retained competence self-assembly into ENCs. To provide cellular specificity, PreS121–47 hepatocyte-targeting peptide, genetically protein, conjugated both versions MxENC. underwent comprehensive characterization stability, cargo loading, uptake, release HepG2 cells, demonstrating their potential as protein-delivery vehicles. These results valuable insights design customization nanocompartments, opening up possibilities improved drug delivery applications biotechnology nanomedicine.

Language: Английский

Citations

0
Structural and Biochemical Characterization of a Widespread Enterobacterial Peroxidase Encapsulin DOI Creative Commons
Natalia C. Ubilla‐Rodriguez, Michael P. Andreas, Tobias W. Giessen

et al.

Advanced Science, Journal Year: 2025, Volume and Issue: unknown

Published: April 1, 2025

Encapsulins are self-assembling protein compartments found in prokaryotes and specifically encapsulate dedicated cargo enzymes. The most abundant encapsulin class Dye-decolorizing Peroxidases (DyPs). It has been previously suggested that DyP encapsulins involved oxidative stress resistance bacterial pathogenicity due to DyPs' inherent ability reduce detoxify hydrogen peroxide while oxidizing a broad range of organic co-substrates. Here, we report the structural biochemical analysis widely across enterobacteria. Using bioinformatic approaches, show this is encoded by conserved transposon-associated operon, enriched enterobacterial pathogens. Through low pH exposure experiments, highlight stability under harsh conditions catalytic activity highest at pH. We determine structure DyP-loaded shell free via cryo-electron microscopy, revealing basis for loading preference. This work lays foundation further explore substrate physiological functions encapsulins.

Language: Английский

Citations

0
Structural and biochemical characterization of a widespread enterobacterial peroxidase encapsulin DOI Creative Commons
Natalia C. Ubilla‐Rodriguez, Michael P. Andreas, Tobias W. Giessen

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Nov. 27, 2024

Encapsulins are self-assembling protein compartments found in prokaryotes and specifically encapsulate dedicated cargo enzymes. The most abundant encapsulin class Dye-decolorizing Peroxidases (DyPs). It has been previously suggested that DyP encapsulins involved oxidative stress resistance bacterial pathogenicity due to DyPs' inherent ability reduce detoxify hydrogen peroxide while oxidizing a broad range of organic co-substrates. Here, we report the structural biochemical analysis widely across enterobacteria. Using bioinformatic approaches, show this is encoded by conserved transposon-associated operon, enriched enterobacterial pathogens. Through low pH exposure experiments, highlight stability under harsh conditions catalytic activity highest at pH. We determine structure DyP-loaded shell free via cryo-electron microscopy, revealing basis for loading preference. Our work lays foundation further explore substrate physiological functions encapsulins.

Language: Английский

Citations

0
Identification and cryoEM Structure Determination of Escherichia Phage YDC107 Tail Found in a Bacteria-Contaminated Buffer DOI Creative Commons
Matthew C. Jenkins, Tapati Dutta, Daija Bobe

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Dec. 11, 2024

Abstract Cryo-electron microscopy data analysis can yield multiple structures from a single heterogeneous dataset. Here, we show workflow used for the identification of contaminant cryoEM grid without prior knowledge protein sequence. We determined tail structure Escherichia phage YDC107 only several thousand particles. The combines high-resolution single-particle processing with de novo model determination using ML-based methods. Structural revealed that central part has C6 symmetry, however overall symmetry each segment is C3 due to dimerization flexible domain. Figure

Language: Английский

Citations

0