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Journal of Bacteriology, February 2008, p. 1350-1358, Vol. 190, No. 4
0021-9193/08/$08.00+0     doi:10.1128/JB.01149-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Xylanase Attachment to the Cell Wall of the Hyperthermophilic Bacterium Thermotoga maritima{triangledown}

Wolfgang Liebl,1* Christoph Winterhalter,2 Wolfgang Baumeister,3 Martin Armbrecht,1,{dagger} and Michael Valdez1

Institut für Mikrobiologie und Genetik, Georg-August-Universität, Grisebachstr. 8, D-37077 Göttingen, Germany,1 Wacker Chemie GmbH, Johannes Hess Str. 24, D-84489 Burghausen, Germany,2 Max-Planck-Institut für Biochemie, Am Klopferspitz, D-82152 Martinsried, Germany3

Received 20 July 2007/ Accepted 27 November 2007

The cellular localization and processing of the endo-xylanases (1,4-β-D-xylan-xylanohydrolase; EC 3.2.1.8) of the hyperthermophile Thermotoga maritima were investigated, in particular with respect to the unusual outer membrane ("toga") of this gram-negative bacterium. XynB (40 kDa) was detected in the periplasmic fraction of T. maritima cells and in the culture supernatant. XynA (120 kDa) was partially released to the surrounding medium, but most XynA remained cell associated. Immunogold labeling of thin sections revealed that cell-bound XynA was localized mainly in the outer membranes of T. maritima cells. Amino-terminal sequencing of purified membrane-bound XynA revealed processing of the signal peptide after the eighth residue, thereby leaving the hydrophobic core of the signal peptide attached to the enzyme. This mode of processing is reminiscent of type IV prepilin signal peptide cleavage. Removal of the entire XynA signal peptide was necessary for release from the cell because enzyme purified from the culture supernatant lacked 44 residues at the N terminus, including the hydrophobic part of the signal peptide. We conclude that toga association of XynA is mediated by residues 9 to 44 of the signal peptide. The biochemical and electron microscopic localization studies together with the amino-terminal processing data indicate that XynA is held at the cell surface of T. maritima via a hydrophobic peptide anchor, which is highly unusual for an outer membrane protein.

* Corresponding author. Mailing address: Institut für Mikrobiologie und Genetik, Grisebachstr. 8, D-37077 Göttingen, Germany. Phone: 49-551-393795. Fax: 49-551-394897. E-mail: wliebl{at}gwdg.de

{triangledown} Published ahead of print on 14 December 2007.

{dagger} Present address: Eppendorf AG, Barkhausenweg 1, D-22339 Hamburg, Germany.

Journal of Bacteriology, February 2008, p. 1350-1358, Vol. 190, No. 4
0021-9193/08/$08.00+0     doi:10.1128/JB.01149-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.





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