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Titel: N-Acetylmuramic Acid (MurNAc) Auxotrophy of the Oral Pathogen Tannerella forsythia: Characterization of a MurNAc Kinase and Analysis of Its Role in Cell Wall Metabolism
VerfasserIn: Hottmann, Isabel
Mayer, Valentina M. T.
Tomek, Markus B.
Friedrich, Valentin
Calvert, Matthew B.
Titz, Alexander
Schäffer, Christina
Mayer, Christoph
Sprache: Englisch
Titel: Frontiers in Microbiology
Bandnummer: 9
Verlag/Plattform: Frontiers
Erscheinungsjahr: 2018
Freie Schlagwörter: oral pathogen
red complex consortium
N-acetylmuramic acid kinase
MurNAc auxotrophy
peptidoglycan metabolism
cell wall recycling
DDC-Sachgruppe: 500 Naturwissenschaften
Dokumenttyp: Journalartikel / Zeitschriftenartikel
Abstract: Tannerella forsythia is an anaerobic, Gram-negative oral pathogen that thrives in multispecies gingival biofilms associated with periodontitis. The bacterium is auxotrophic for the commonly essential bacterial cell wall sugar N-acetylmuramic acid (MurNAc) and, thus, strictly depends on an exogenous supply of MurNAc for growth and maintenance of cell morphology. A MurNAc transporter (Tf_MurT; Tanf_08375) and an ortholog of the Escherichia coli etherase MurQ (Tf_MurQ; Tanf_08385) converting MurNAc-6-phosphate to GlcNAc-6-phosphate were recently described for T. forsythia. In between the respective genes on the T. forsythia genome, a putative kinase gene is located. In this study, the putative kinase (Tf_MurK; Tanf_08380) was produced as a recombinant protein and biochemically characterized. Kinetic studies revealed Tf_MurK to be a 6-kinase with stringent substrate specificity for MurNAc exhibiting a 6 × 104 - fold higher catalytic efficiency (kcat/Km) for MurNAc than for N-acetylglucosamine (GlcNAc) with kcat values of 10.5 s−1 and 0.1 s−1 and Km values of 200 µM and 116 mM, respectively. The enzyme kinetic data suggest that Tf_MurK is subject to substrate inhibition (Ki[S] = 4.2 mM). To assess the role of Tf_MurK in the cell wall metabolism of T. forsythia, a kinase deletion mutant (1Tf_murK::erm) was constructed. This mutant accumulated MurNAc intracellularly in the exponential phase, indicating the capability to take up MurNAc, but inability to catabolize MurNAc. In the stationary phase, the MurNAc level was reduced in the mutant, while the level of the peptidoglycan precursor UDP-MurNAc-pentapeptide was highly elevated. Further, according to scanning electron microscopy evidence, the 1Tf_murK::erm mutant was more tolerant toward low MurNAc concentration in the medium (below 0.5 µg/ml) before transition from healthy, rod-shaped to fusiform cells occurred, while the parent strain required > 1 µg/ml MurNAc for optimal growth. These data reveal that T. forsythia readily catabolizes exogenous MurNAc but simultaneously channels a proportion of the sugar into peptidoglycan biosynthesis. Deletion of Tf_murK blocks MurNAc catabolism and allows the direction of MurNAc solely to peptidoglycan biosynthesis, resulting in a growth advantage in MurNAc-depleted medium. This work increases our understanding of the T. forsythia cell wall metabolism and may pave new routes for lead finding in the treatment of periodontitis.
DOI der Erstveröffentlichung: 10.3389/fmicb.2018.00019
URL der Erstveröffentlichung: https://doi.org/10.3389/fmicb.2018.00019
Link zu diesem Datensatz: urn:nbn:de:bsz:291--ds-392431
hdl:20.500.11880/35371
http://dx.doi.org/10.22028/D291-39243
ISSN: 1664-302X
Datum des Eintrags: 7-Mär-2023
Bezeichnung des in Beziehung stehenden Objekts: Supplementary Material
In Beziehung stehendes Objekt: https://www.frontiersin.org/articles/file/downloadfile/331701_supplementary-materials_datasheets_1_docx/octet-stream/Data%20Sheet%201.DOCX/1/331701
Fakultät: NT - Naturwissenschaftlich- Technische Fakultät
Fachrichtung: NT - Chemie
Professur: NT - Univ.-Prof. Dr. phil. Alexander Titz
Sammlung:SciDok - Der Wissenschaftsserver der Universität des Saarlandes

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