Gp29 LysA of mycobacteriophage TM4 can hydrolyze peptidoglycan through an N-acetyl-muramoyl-L-alanine amidase activity

Autor: Estefanía Urdániz, Mariano Martín, Florencia Payaslián, Lucas Alfredo Defelipe, Martín Dodes, Mariano Martinez, Pedro M. Alzari, Gabriela Cabrera, Marcelo Adrián Martí, Mariana Piuri
Přispěvatelé: Facultad de Ciencias Exactas y Naturales [Buenos Aires] (FCEyN), Universidad de Buenos Aires [Buenos Aires] (UBA), Universidad Nacional de Córdoba [Argentina], European Molecular Biology Laboratory [Hamburg] (EMBL), Microbiologie structurale - Structural Microbiology (Microb. Struc. (UMR_3528 / U-Pasteur_5)), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité), This research was supported by Agencia Nacional de Promoción Científica y Tecnológica (PIDC-0015/2015) to MP. MM and FP are CONICET fellows. LAD was funded by EMBL Interdisciplinary Postdoc Programme under Marie Curie Actions COFUND 664726., European Project: 664726,H2020,H2020-MSCA-COFUND-2014,EI3POD(2015)
Jazyk: angličtina
Rok vydání: 2022
Předmět:
Endolysin
[SDV]Life Sciences [q-bio]
Mycobacterium smegmatis
MESH: Micrococcus
Biophysics
N-acetyl-muramoyl-L-alanine amidase
Peptidoglycan
Galactans
Biochemistry
Mass Spectrometry
MESH: Muramic Acids
Micrococcus
Analytical Chemistry
Viral Proteins
LysA
MESH: Mycobacteriophages
Endopeptidases
Escherichia coli
[SDV.BBM]Life Sciences [q-bio]/Biochemistry
Molecular Biology
MESH: Endopeptidases
Molecular Biology
MESH: Mycobacterium smegmatis
MESH: Mass Spectrometry
Mycobacteriophage
MESH: Peptidoglycan
MESH: Escherichia coli
Hydrolysis
Computational Biology
MESH: Galactans
Mycobacteriophages
N-Acetylmuramoyl-L-alanine Amidase
MESH: Viral Proteins
[SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology
Muramic Acids
MESH: N-Acetylmuramoyl-L-alanine Amidase
[INFO.INFO-BI]Computer Science [cs]/Bioinformatics [q-bio.QM]
MESH: Hydrolysis
MESH: Computational Biology
Zdroj: Biochimica et Biophysica Acta (BBA)-Proteins and Proteomics
Biochimica et Biophysica Acta (BBA)-Proteins and Proteomics, 2022, 1870 (2), pp.140745. ⟨10.1016/j.bbapap.2021.140745⟩
ISSN: 1570-9639
Popis: International audience; Bacteriophage endolysins are crucial for progeny release at the end of the lytic cycle. Mycobacteriophage's genomes carry a lysin A essential gene, whose product cleaves the peptidoglycan (PG) layer and a lysin B, coding for an esterase, that cleaves the linkage between the mycolic acids and the arabinogalactan-PG complex. Lysin A mycobacteriophage proteins are highly modular and in gp29 (LysA) of phage TM4 three distinctive domains were identified. By bioinformatics analysis the central module was previously found to be similar to an amidase-2 domain family with an N-acetylmuramoyl-L-alanine amidase activity. We demonstrated experimentally that purified LysA is able to lyse a suspension of Micrococcus lysodeikticus and can promote cell lysis when expressed in E. coli and Mycobacterium smegmatis. After incubation of LysA with MDP (Muramyl dipeptide, N-acetyl-muramyl-L-alanyl-D-isoglutamine) we detected the presence of N-acetylmuramic acid (NAcMur) and L-Ala-D-isoGlutamine (L-Ala-D-isoGln) corroborating the proposed muramidase activity of this enzyme. This protein was stabilized at acidic pH in the presence of Zn consistent with the increase of the enzymatic activity under these conditions. By homology modeling, we predicted that the Zn ion is coordinated by His 226, His 335, and Asp 347 and we also identified the amino acid Glu 290 as the catalytic residue. LysA activity was completely abolished in derived mutants on these key residues, suggesting that the PG hydrolysis solely relies on the central domain of the protein.
Databáze: OpenAIRE
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