Internal cell-penetrating peptide-mediated internalization enables a chimeric lysin to target intracellular pathogens

Yang, Hang; Xu, Jingjing; Gong, Yujing; Tang, Yan; Li, Wuyou; Zheng, Zhenhua; Li, Yuhong; He, Jin; Wei, Hongping

doi:10.1016/j.ijpharm.2021.120449

Cited by 5 publications

(6 citation statements)

References 38 publications

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“…Additionally, we identified an optimal position of this TAT at the N-terminus, while the cited studies all rationally fused this specific CPP C-terminally. For completion, it can be remarked that the PlySs2 SH3_5 and PlyCb domains included in this study have also been attributed certain CPP functionalities (Shen et al, 2016;Yang et al, 2021).…”

Section: Discussionmentioning

confidence: 87%

“…This might be due to several differences between ClyR and our study such as screening conditions and differences in delineation of the subdomains (Röhrig et al, 2020 ; Vander Elst et al, 2020 ). Notably, follow‐up studies with ClyR only focused on streptococcal strains (Xu et al, 2018 ; Yang et al, 2016 , 2021 ; Zhu et al, 2021 ).…”

Section: Discussionmentioning

confidence: 99%

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Development of engineered endolysins with in vitro intracellular activity against streptococcal bovine mastitis‐causing pathogens

Vander Elst,

Bert,

Favoreel

et al. 2023

Microbial Biotechnology

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Bacteriophage‐derived endolysins are a novel class of antimicrobials known to rapidly kill bacteria, including antibiotic‐resistant strains. We here engineered endolysins against the bovine mastitis pathogens Streptococcus uberis, Streptococcus agalactiae and Streptococcus dysgalactiae, also targeting intracellular survival and biofilm formation. For this purpose, high‐throughput DNA assembly was used to create a library with >80,000 theoretical endolysin variants for screening of their bacteriolytic activity against Gram‐positive isolates from (sub)clinically affected cows. This lytic activity was evaluated by turbidity reduction and time‐kill assays in phosphate‐buffered saline and pasteurized whole cow's milk to allow a rank up of the most potent leading candidates. A top candidate was selected with a 4.0 log killing efficacy against S. uberis, also showing similar activity against S. agalactiae and S. dysgalactiae. This top candidate eradicated S. uberis biofilm and showed intracellular activity in two bovine mammary epithelial cell lines as was confirmed by confocal microscopy. A potentiating effect on cloxacillin, a beta‐lactam penicillin used to intramammarily treat bovine Gram‐positive mastitis, was observed for this top candidate endolysin in raw cow's milk from (sub)clinically infected udders. Our in vitro results indicate that engineered endolysins may have a future role as add‐on in the treatment of bovine streptococcal mastitis.

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Section: Discussionmentioning

confidence: 87%

Section: Discussionmentioning

confidence: 99%

Development of engineered endolysins with in vitro intracellular activity against streptococcal bovine mastitis‐causing pathogens

Vander Elst,

Bert,

Favoreel

et al. 2023

Microbial Biotechnology

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“…The authors state that this is not a surprising finding, but rather underscores the importance of the EAD to be present for full binding functionality of PlySs2 [ 16 ]. For completion, PlySs2 SH3_5 has also been attributed several eukaryotic cell penetrating properties [ 76 ]. To proceed, PlySs2 has been characterized with potent biofilm eradicating activity [ 77 , 78 ].…”

Section: Reviewmentioning

confidence: 99%

“…, additional ΔOD 600nm in the range of ± 0.15–0.35 for 3 S. aureus isolates). It has also been shown that ClyR is able to tackle streptococci intracellularly, due to certain cell penetrating properties of PlySs2 SH3_5 [ 76 ]. The intracellular uptake of ClyR is believed to happen through caveolin-dependent endocytosis and was shown by confocal laser microscopy, after exposing human adenocarcinoma A549 cells with 100 µg/mL Alexa Fluor 488-labeled ClyR during 40 min.…”

Section: Reviewmentioning

confidence: 99%

Bacteriophage-derived endolysins as innovative antimicrobials against bovine mastitis-causing streptococci and staphylococci: a state-of-the-art review

Vander Elst

2024

Acta Vet Scand

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Bacteriophage-encoded endolysins, peptidoglycan hydrolases breaking down the Gram-positive bacterial cell wall, represent a groundbreaking class of novel antimicrobials to revolutionize the veterinary medicine field. Wild-type endolysins exhibit a modular structure, consisting of enzymatically active and cell wall-binding domains, that enable genetic engineering strategies for the creation of chimeric fusion proteins or so-called ‘engineered endolysins’. This biotechnological approach has yielded variants with modified lytic spectrums, introducing new possibilities in antimicrobial development. However, the discovery of highly similar endolysins by different groups has occasionally resulted in the assignment of different names that complicate a straightforward comparison. The aim of this review was to perform a homology-based comparison of the wild-type and engineered endolysins that have been characterized in the context of bovine mastitis-causing streptococci and staphylococci, grouping homologous endolysins with ≥ 95.0% protein sequence similarity. Literature is explored by homologous groups for the wild-type endolysins, followed by a chronological examination of engineered endolysins according to their year of publication. This review concludes that the wild-type endolysins encountered persistent challenges in raw milk and in vivo settings, causing a notable shift in the field towards the engineering of endolysins. Lead candidates that display robust lytic activity are nowadays selected from screening assays that are performed under these challenging conditions, often utilizing advanced high-throughput protein engineering methods. Overall, these recent advancements suggest that endolysins will integrate into the antibiotic arsenal over the next decade, thereby innovating antimicrobial treatment against bovine mastitis-causing streptococci and staphylococci.

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“…Moreover, Researchers have attempted to expand the endolysin customization possibilities by fusing it with functional domains. For example, linking endolysin with an albumin‐binding domain to increase its half‐life in blood or fusing it with cell‐penetrating peptides to enable endolysin to enter mammalian cells and kill intracellular bacteria (Seijsing et al., 2018 ; Yang et al., 2021 ).…”

Section: Strategy Of Endolysin Against Gram‐negative Bacteriamentioning

confidence: 99%

Engineering strategies and challenges of endolysin as an antibacterial agent against Gram‐negative bacteria

Zheng,

Zhang

2024

Microbial Biotechnology

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Bacteriophage endolysin is a novel antibacterial agent that has attracted much attention in the prevention and control of drug‐resistant bacteria due to its unique mechanism of hydrolysing peptidoglycans. Although endolysin exhibits excellent bactericidal effects on Gram‐positive bacteria, the presence of the outer membrane of Gram‐negative bacteria makes it difficult to lyse them extracellularly, thus limiting their application field. To enhance the extracellular activity of endolysin and facilitate its crossing through the outer membrane of Gram‐negative bacteria, researchers have adopted physical, chemical, and molecular methods. This review summarizes the characterization of endolysin targeting Gram‐negative bacteria, strategies for endolysin modification, and the challenges and future of engineering endolysin against Gram‐negative bacteria in clinical applications, to promote the application of endolysin in the prevention and control of Gram‐negative bacteria.

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Internal cell-penetrating peptide-mediated internalization enables a chimeric lysin to target intracellular pathogens

Cited by 5 publications

References 38 publications

Development of engineered endolysins with in vitro intracellular activity against streptococcal bovine mastitis‐causing pathogens

Development of engineered endolysins with in vitro intracellular activity against streptococcal bovine mastitis‐causing pathogens

Bacteriophage-derived endolysins as innovative antimicrobials against bovine mastitis-causing streptococci and staphylococci: a state-of-the-art review

Engineering strategies and challenges of endolysin as an antibacterial agent against Gram‐negative bacteria

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