Structure and function of the Ts2631 endolysin of Thermus scotoductus phage vB_Tsc2631 with unique N-terminal extension used for peptidoglycan binding

Magdalena Płotka , Enea Sancho-Vaello , Sebastian Dorawa , Anna Kaczorowska , Lukasz P. Kozlowski , Tadeusz Kaczorowski , Kornelius Zeth

Abstract

To escape from hosts after completing their life cycle, bacteriophages often use endolysins, which degrade bacterial peptidoglycan. While mesophilic phages have been extensively studied, their thermophilic counterparts are not well characterized. Here, we present a detailed analysis of the structure and function of Ts2631 endolysin from thermophilic phage vB_Tsc2631, which is a zincdependent amidase. The active site of Ts2631 consists of His30, Tyr58, His131 and Cys139, which are involved in Zn2+ coordination and catalysis. We found that the active site residues are necessary for lysis yet not crucial for peptidoglycan binding. To elucidate residues involved in the enzyme interaction with peptidoglycan, we tested single-residue substitution variants and identified Tyr60 and Lys70 as essential residues. Moreover, substitution of Cys80, abrogating disulfide bridge formation, inactivates Ts2631, as do substitutions of His31, Thr32 and Asn85 residues. The endolysin contains a positively charged N-terminal extension of 20 residues that can protrude from the remainder of the enzyme and is crucial for peptidoglycan binding. We show that the deletion of 20 residues from the N-terminus abolished the bacteriolytic activity of the enzyme. Because Ts2631 exhibits intrinsic antibacterial activity and unusual thermal stability, it is perfectly suited as a scaffold for the development of antimicrobial agents.
Author Magdalena Płotka (FB / DM)
Magdalena Płotka,,
- Department of Microbiology
, Enea Sancho-Vaello
Enea Sancho-Vaello,,
-
, Sebastian Dorawa (FB / DM)
Sebastian Dorawa,,
- Department of Microbiology
, Anna Kaczorowska (FB / CPM)
Anna Kaczorowska,,
- Collection of Plasmids and Microorganisms
, Lukasz P. Kozlowski
Lukasz P. Kozlowski,,
-
, Tadeusz Kaczorowski (FB / DM)
Tadeusz Kaczorowski,,
- Department of Microbiology
, Kornelius Zeth
Kornelius Zeth,,
-
Journal seriesScientific Reports, ISSN 2045-2322, (A 40 pkt)
Issue year2019
Vol9
Pages1-14
Publication size in sheets0.65
Keywords in Englishcell lysis, amidase, thermal stability, site directed mutagenesis, catalytic center
ASJC Classification1000 Multidisciplinary
DOIDOI:10.1038/s41598-018-37417-6
URL https://doi.org/10.1038/s41598-018-37417-6
Languageen angielski
LicenseJournal (articles only); published final; Uznanie Autorstwa (CC-BY); with publication
Score (nominal)40
ScoreMinisterial score = 40.0, 04-04-2019, ArticleFromJournal
Ministerial score (2013-2016) = 40.0, 04-04-2019, ArticleFromJournal
Publication indicators WoS Citations = 0; Scopus SNIP (Source Normalised Impact per Paper): 2016 = 1.401; WoS Impact Factor: 2017 = 4.122 (2) - 2017=4.609 (5)
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