TY - JOUR A1 - Ribitsch, D. A1 - Karl, W. A1 - Birner-Gruenberger, R. A1 - Gruber, K. A1 - Eiteljoerg, I. A1 - Remler, P. A1 - Wieland, S. A1 - Siegert, Petra A1 - Maurer, Karl-Heinz A1 - Schwab, H. T1 - C-terminal truncation of a metagenome-derived detergent protease for effective expression in E. coli JF - Journal of biotechnology N2 - Recently, a new alkaline protease named HP70 showing highest homology to extracellular serine proteases of Stenotrophomonas maltophilia and Xanthomonas campestris was found in the course of a metagenome screening for detergent proteases (Niehaus et al., submitted for publication). Attempts to efficiently express the enzyme in common expression hosts had failed. This study reports on the realization of overexpression in Escherichia coli after structural modification of HP70. Modelling of HP70 resulted in a two-domain structure, comprising the catalytic domain and a C-terminal domain which includes about 100 amino acids. On the basis of the modelled structure the enzyme was truncated by deletion of most of the C-terminal domain yielding HP70-C477. This structural modification allowed effective expression of active enzyme using E. coli BL21-Gold as the host. Specific activity of HP70-C477 determined with suc-l-Ala-l-Ala-l-Pro-l-Phe-p-nitroanilide as the substrate was 30 ± 5 U/mg compared to 8 ± 1 U/mg of the native enzyme. HP70-C477 was most active at 40 °C and pH 7–11; these conditions are prerequisite for a potential application as detergent enzyme. Determination of kinetic parameters at 40 °C and pH = 9.5 resulted in KM = 0.23 ± 0.01 mM and kcat = 167.5 ± 3.6 s⁻¹. MS-analysis of peptide fragments obtained from incubation of HP70 and HP70-C477 with insulin B indicated that the C-terminal domain influences the cleavage preferences of the enzyme. Washing experiments confirmed the high potential of HP70-C477 as detergent protease. Y1 - 2010 U6 - https://doi.org/10.1016/j.jbiotec.2010.09.947 SN - 1873-4863 (E-Journal); 0168-1656 (Print) VL - 150 IS - 3 SP - 408 EP - 416 PB - Elsevier CY - Amsterdam ER - TY - PAT A1 - Siegert, Petra A1 - Mussmann, Nina A1 - O'Connell, Timothy A1 - Maurer, Karl-Heinz T1 - Neue Proteasen und Mittel enthaltend diese Proteasen [Offenlegungsschrift] T1 - Novel proteases and means containing said proteases [Internationale Patentanmeldung] Y1 - 2010 SP - 1 EP - 30 PB - Deutsches Patent- und Markenamt / WIPO CY - München / Genf ER - TY - PAT A1 - Siegert, Petra A1 - Baumstark, Rebecca A1 - Kluin, Cornelia A1 - O'Connell, Timothy A1 - Maurer, Karl-Heinz A1 - Hellmuth, Hendrik T1 - Neue Proteasen und Mittel enthaltend diese Proteasen [Offenlegungsschrift] Y1 - 2010 SP - 1 EP - 30 PB - Deutsches Patent- und Markenamt CY - München ER - TY - PAT A1 - Siegert, Petra A1 - Spitz, Astrid A1 - Maurer, Karl-Heinz T1 - Neue Proteasen und Mittel enthaltend diese Proteasen [Offenlegungsschrift] T1 - Novel proteases and compositions comprising these proteases [Internationale Patentanmeldung] Y1 - 2010 SP - 1 EP - 31 PB - Deutsches Patentamt / WIPO CY - München / Genf ER - TY - PAT A1 - Siegert, Petra A1 - Spitz, Astrid A1 - Maurer, Karl-Heinz T1 - Wasch- und Reinigungsmittel enthaltend Proteasen aus Bacillus pumilus [Offenlegungsschrift] T1 - Detergents and cleaning agents containing proteases from bacillus pumilus [Europäische Patentschrift / Internationale Patentanmeldung] Y1 - 2010 SP - 1 EP - 20 PB - Deutsches Patentamt / Europäisches Patentamt / WIPO CY - München / Den Hague / Genf ER - TY - CHAP A1 - Buda, Aurel A1 - Schürmann, Volker A1 - Wollert, Jörg T1 - Wireless technologies in factory automation T2 - Factory automation / ed. by Javier Silvestre-Blanes Y1 - 2010 SN - 978-953-7619-42-8 SP - 29 EP - 50 PB - Intech CY - London ER - TY - JOUR A1 - Wollert, Jörg A1 - Leidinger, Rafael T1 - Automation trifft Automotive : SPS in der Fahrzeugtechnik JF - Elektronik automotive : Magazin für Entwicklungen in der Kfz-Elektronik und Telematik N2 - Eine SPS im Kraftfahrzeug? Das geht ja gar nicht – da gibt es doch all die Hersteller für Kfz-Steuergeräte, die konfigurierbare Hardware anbieten. Im Prinzip ja, aber leider sind gerade die Sensoren oder Aktoren nicht ansteuerbar, die der Entwickler genau für sein aktuelles Projekt benötigt. Dieser Beitrag gibt einen kleinen Einblick in neue Möglichkeiten des Rapid Prototyping für mechatronische Systeme auf der Basis von Speicherprogrammierbaren Steuerungen (SPS). Y1 - 2010 SN - 1614-0125 IS - 4/5 SP - 44 EP - 48 PB - WEKA Fachmedien CY - Haar ER - TY - CHAP A1 - Schürmann, Volker A1 - Keller, A. A1 - Wollert, Jörg T1 - Gebäudeautomation mit Webtechnologien : ein Erfahrungsbericht T2 - Automation 2010 : der 11. Branchentreff der Mess- und Automatisierungstechnik ; Kongress Baden-Baden, 15. und 16. Juni 2010. (VDI-Berichte ; 2092) Y1 - 2010 SN - 978-3-18-092092-4 SP - 321 EP - 324 PB - VDI-Verl. CY - Düsseldorf ER - TY - CHAP A1 - Schürmann, Volker A1 - Buda, Aurel A1 - Jonker, Stefan A1 - Palmhof, Norman A1 - Wollert, Jörg T1 - Wireless in future automotive applications T2 - Mobile and wireless communications network layer and circuit level design / ed. by Salma Ait Fares ... Y1 - 2010 SN - 978-953-307-042-1 SP - 71 EP - 92 PB - Intech CY - London ER - TY - CHAP A1 - Schürmann, Volker A1 - Leidinger, R. A1 - Wollert, Jörg T1 - Simplified rapid-prototyping with automation technology T2 - Automobiles and sustainable mobility : FISITA 2010 World Automotive Congress, 30 May - 4 June, Budapest, Hungary ; proceedings Y1 - 2010 SN - 978-963-9058-29-3 SP - F2010C093 ER -