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Journal of Bacteriology, July 2009, p. 4056-4069, Vol. 191, No. 13
0021-9193/09/$08.00+0     doi:10.1128/JB.00117-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Transcriptome and Functional Analysis of the Eukaryotic-Type Serine/Threonine Kinase PknB in Staphylococcus aureus{triangledown}

Stefanie Donat,1 Karin Streker,1 Tanja Schirmeister,2 Sonja Rakette,3 Thilo Stehle,3 Manuel Liebeke,4 Michael Lalk,4 and Knut Ohlsen1*

Universität Würzburg, Institut für Molekulare Infektionsbiologie, Würzburg, Germany,1 Universität Würzburg, Institut für Pharmazie und Lebensmittelchemie, Würzburg, Germany,2 Universität Tübingen, Interfakultäres Institut für Biochemie, Strukturbiologie, Tübingen, Germany,3 Ernst-Moritz-Arndt-Universität Greifswald, Institut für Pharmazie, Kompetenzzentrum Functional Genomics, Greifswald, Germany4

Received 28 January 2009/ Accepted 6 April 2009

The function of the Staphylococcus aureus eukaryotic-like serine/threonine protein kinase PknB was investigated by performing transcriptome analysis using DNA microarray technology and biochemical assays. The transcriptional profile revealed a strong regulatory impact of PknB on the expression of genes encoding proteins which are involved in purine and pyrimidine biosynthesis, cell wall metabolism, autolysis, and glutamine synthesis. Functional activity of overexpressed and purified PknB kinase was demonstrated using the myelin basic protein as a surrogate substrate. Phosphorylation occurred in a time-dependent manner with Mn2+ as a preferred cofactor. Furthermore, biochemical characterization revealed regulation of adenylosuccinate synthase (PurA) activity by phosphorylation. Phosphorylated PurA showed a 1.8-fold decrease in enzymatic activity compared to unphosphorylated PurA. Loss of PknB led to formation of larger cell clusters, and a pknB deletion strain showed 32-fold-higher sensitivity to the cell wall-active antibiotic tunicamycin. The results of this study strongly indicate that PknB has a role in regulation of purine biosynthesis, autolysis, and central metabolic processes in S. aureus.

* Corresponding author. Mailing address: Institut für Molekulare Infektionsbiologie, Röntgenring 11, D-97070 Würzburg, Germany. Phone: 49-931-312155. Fax: 49-931-312578. E-mail: knut.ohlsen{at}mail.uni-wuerzburg.de

{triangledown} Published ahead of print on 17 April 2009.

Journal of Bacteriology, July 2009, p. 4056-4069, Vol. 191, No. 13
0021-9193/09/$08.00+0     doi:10.1128/JB.00117-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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