Metaviromics analysis of marine biofilm reveals a glycoside hydrolase endolysin with high specificity towards Acinetobacter baumannii

Premetis, Georgios; Georgakis, Nikolaos; Stathi, Angeliki; Labrou, Nikolaos E.

doi:10.1016/j.bbapap.2023.140918

Cited by 5 publications

(3 citation statements)

References 52 publications

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“…As can be observed, the melting temperature of all mentioned endolysins differs only by a few degrees and is in a range of 91.70°C-103.56°C. For comparison, the newly discovered AbLys2 endolysin from metavirome of marine biofilm sample that shows lytic activity against mesophilic bacteria A. baumannii has a T m of 47.1°C (Premetis et al, 2023). The addition of EDTA to PhiKo endolysin decreases the T m to 86.0°C (Figure 5B).…”

Section: Discussionmentioning

confidence: 99%

Molecular characterization of the PhiKo endolysin from Thermus thermophilus HB27 bacteriophage phiKo and its cryptic lytic peptide RAP-29

Szadkowska,

Kocot,

Sowik

et al. 2024

Front. Microbiol.

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IntroductionIn the era of increasing bacterial resistance to antibiotics, new bactericidal substances are sought, and lysins derived from extremophilic organisms have the undoubted advantage of being stable under harsh environmental conditions. The PhiKo endolysin is derived from the phiKo bacteriophage infecting Gram-negative extremophilic bacterium Thermus thermophilus HB27. This enzyme shows similarity to two previously investigated thermostable type-2 amidases, the Ts2631 and Ph2119 from Thermus scotoductus bacteriophages, that revealed high lytic activity not only against thermophiles but also against Gram-negative mesophilic bacteria. Therefore, antibacterial potential of the PhiKo endolysin was investigated in the study presented here.MethodsEnzyme activity was assessed using turbidity reduction assays (TRAs) and antibacterial tests. Differential scanning calorimetry was applied to evaluate protein stability. The Collection of Anti-Microbial Peptides (CAMP) and Antimicrobial Peptide Calculator and Predictor (APD3) were used to predict regions with antimicrobial potential in the PhiKo primary sequence. The minimum inhibitory concentration (MIC) of the RAP-29 synthetic peptide was determined against Gram-positive and Gram-negative selected strains, and mechanism of action was investigated with use of membrane potential sensitive fluorescent dye 3,3′-Dipropylthiacarbocyanine iodide (DiSC3(5)).Results and discussionThe PhiKo endolysin is highly thermostable with melting temperature of 91.70°C. However, despite its lytic effect against such extremophiles as: T. thermophilus, Thermus flavus, Thermus parvatiensis, Thermus scotoductus, and Deinococcus radiodurans, PhiKo showed moderate antibacterial activity against mesophiles. Consequently, its protein sequence was searched for regions with potential antibacterial activity. A highly positively charged region was identified and synthetized (PhiKo105-133). The novel RAP-29 peptide lysed mesophilic strains of staphylococci and Gram-negative bacteria, reducing the number of cells by 3.7–7.1 log units and reaching the minimum inhibitory concentration values in the range of 2–31 μM. This peptide is unstructured in an aqueous solution but forms an α-helix in the presence of detergents. Moreover, it binds lipoteichoic acid and lipopolysaccharide, and causes depolarization of bacterial membranes. The RAP-29 peptide is a promising candidate for combating bacterial pathogens. The existence of this cryptic peptide testifies to a much wider panel of antimicrobial peptides than thought previously.

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Section: Discussionmentioning

confidence: 99%

Molecular characterization of the PhiKo endolysin from Thermus thermophilus HB27 bacteriophage phiKo and its cryptic lytic peptide RAP-29

Szadkowska,

Kocot,

Sowik

et al. 2024

Front. Microbiol.

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“…Endolysins or PG hydrolases constitute a specific group of enzybiotics encoded by bacteriophages and/or prophages in host cells at the end of the bacteriophage lytic cycle (Murray et al 2021 ; Love et al 2020 ). Previous studies have proven that endolysins exhibit lytic activity against the majority of resistant bacterial cells (Loeffler et al 2003 ; Harhala et al 2018 ; Abdelkader et al 2019 ; Dams and Briers 2019 ; Premetis et al 2023a , b , c ). Moreover, evaluation tests of endolysins have confirmed their non-toxic behavior after their administration in animal models (Rodríguez-Rubio et al 2013 ; Hermoso et al 2003 ; Kashani et al 2018 ; Guo et al 2017 ).…”

Section: Introductionmentioning

confidence: 99%

“…Hence, they could be termed as the “tropical rainforests” of the microbial realm (Des Marais and Walter 2019 ). Metagenomics enable scientists to obtain genetic resources for novel proteins, enzymes, and metabolites from organisms that are not cultivable, allowing the direct assessment of huge genetic diversity within natural microbial communities, leading to the discovery of novel genes, genomes, and microbial functions (Premetis et al 2023c ). Extremophiles, which are naturally occurring in “extreme” ecological niches, generate thermostable enzymes with economic significance that are useful for bioprocesses and product development (Goh et al 2022 ; Swift et al 2019 ).…”

Section: Introductionmentioning

confidence: 99%

Metagenomic analysis of hot spring soil for mining a novel thermostable enzybiotic

Pantiora,

Georgakis,

Premetis

et al. 2024

Appl Microbiol Biotechnol

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The misuse and overuse of antibiotics have contributed to a rapid emergence of antibiotic-resistant bacterial pathogens. This global health threat underlines the urgent need for innovative and novel antimicrobials. Endolysins derived from bacteriophages or prophages constitute promising new antimicrobials (so-called enzybiotics), exhibiting the ability to break down bacterial peptidoglycan (PG). In the present work, metagenomic analysis of soil samples, collected from thermal springs, allowed the identification of a prophage-derived endolysin that belongs to the N-acetylmuramoyl-L-alanine amidase type 2 (NALAA-2) family and possesses a LysM (lysin motif) region as a cell wall binding domain (CWBD). The enzyme (Ami1) was cloned and expressed in Escherichia coli, and its bactericidal and lytic activity was characterized. The results indicate that Ami1 exhibits strong bactericidal and antimicrobial activity against a broad range of bacterial pathogens, as well as against isolated peptidoglycan (PG). Among the examined bacterial pathogens, Ami1 showed highest bactericidal activity against Staphylococcus aureus sand Staphylococcus epidermidis cells. Thermostability analysis revealed a melting temperature of 64.2 ± 0.6 °C. Overall, these findings support the potential that Ami1, as a broad spectrum antimicrobial agent, could be further assessed as enzybiotic for the effective treatment of bacterial infections. Key points • Metagenomic analysis allowed the identification of a novel prophage endolysin • The endolysin belongs to type 2 amidase family with lysin motif region • The endolysin displays high thermostability and broad bactericidal spectrum

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Mining and multifaceted applications of phage lysin for combatting Vibrio parahaemolyticus

Liu,

Wu,

Malakar

et al. 2024

Food Research International

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Metaviromics analysis of marine biofilm reveals a glycoside hydrolase endolysin with high specificity towards Acinetobacter baumannii

Cited by 5 publications

References 52 publications

Molecular characterization of the PhiKo endolysin from Thermus thermophilus HB27 bacteriophage phiKo and its cryptic lytic peptide RAP-29

Molecular characterization of the PhiKo endolysin from Thermus thermophilus HB27 bacteriophage phiKo and its cryptic lytic peptide RAP-29

Metagenomic analysis of hot spring soil for mining a novel thermostable enzybiotic

Mining and multifaceted applications of phage lysin for combatting Vibrio parahaemolyticus

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