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AmiP from hyperthermophilic Thermus parvatiensis prophage is a thermoactive and ultrathermostable peptidoglycan lytic amidase

Jasilionis, Andrius; Plotka, Magdalena; Wang, Lei; Dorawa, Sebastian; Lange, Joanna; Watzlawick, Hildegard; van den Bergh, Tom; Vroling, Bas; Altenbuchner, Josef; Kaczorowska, Anna‐Karina; Pohl, Ehmke; Kaczorowski, Tadeusz; Nordberg Karlsson, Eva; Freitag‐Pohl, Stefanie

AmiP from hyperthermophilic Thermus parvatiensis prophage is a thermoactive and ultrathermostable peptidoglycan lytic amidase Thumbnail


Authors

Andrius Jasilionis

Magdalena Plotka

Lei Wang

Sebastian Dorawa

Joanna Lange

Hildegard Watzlawick

Tom van den Bergh

Bas Vroling

Josef Altenbuchner

Anna‐Karina Kaczorowska

Tadeusz Kaczorowski

Eva Nordberg Karlsson



Abstract

Bacteriophages encode a wide variety of cell wall disrupting enzymes that aid the viral escape in the final stages of infection. These lytic enzymes have accumulated notable interest due to their potential as novel antibacterials for infection treatment caused by multiple-drug resistant bacteria. Here, the detailed functional and structural characterization of Thermus parvatiensis prophage peptidoglycan lytic amidase AmiP, a globular Amidase_3 type lytic enzyme adapted to high temperatures is presented. The sequence and structure comparison with homologous lytic amidases reveals the key adaptation traits that ensure the activity and stability of AmiP at high temperatures. The crystal structure determined at a resolution of 1.8 Å displays a compact α/β-fold with multiple secondary structure elements omitted or shortened compared to protein structures of similar proteins. The functional characterisation of AmiP demonstrates high efficiency of catalytic activity and broad substrate specificity towards thermophilic and mesophilic bacteria strains containing Orn-type or DAP-type peptidoglycan. The here presented AmiP constitutes the most thermoactive and ultrathermostable Amidase_3 type lytic enzyme biochemically characterised with a temperature optimum at 85 °C. The extraordinary high melting temperature Tm 102.6 °C confirms fold stability up to approximately 100 °C. Furthermore, AmiP is shown to be more active over the alkaline pH range with pH optimum at pH 8.5 and tolerates NaCl up to 300 mM with the activity optimum at 25 mM NaCl. This set of beneficial characteristics suggests that AmiP can be further exploited in biotechnology.

Journal Article Type Article
Acceptance Date Jan 27, 2023
Online Publication Date Jan 31, 2023
Publication Date 2023-03
Deposit Date Feb 6, 2023
Publicly Available Date Mar 29, 2023
Journal Protein Science
Print ISSN 0961-8368
Electronic ISSN 1469-896X
Publisher Wiley
Peer Reviewed Peer Reviewed
Volume 32
Issue 3
Article Number e4585
DOI https://doi.org/10.1002/pro.4585
Public URL https://durham-repository.worktribe.com/output/1180055