TY - PAT A1 - O'Connell, Timothy A1 - Hoven, Nina A1 - Siegert, Petra A1 - Maurer, Karl-Heinz T1 - Amadoriasen in Wasch- und Reinigungsmitteln [Offenlegungsschrift] T1 - Amadoriases in washing and cleaning products [Europäische Patentanmeldung / Internationale Patentanmeldung] Y1 - 2007 SP - 1 EP - 45 PB - Deutsches Patent- und Markenamt / Europäisches Patentamt / WIPO CY - München / Den Hague / Genf ER - 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 - http://dx.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 - JOUR A1 - Niehaus, F. A1 - Gabor, E. A1 - Wieland, S. A1 - Siegert, Petra A1 - Maurer, Karl-Heinz A1 - Eck, J. T1 - Enzymes for the laundry industries: tapping the vast metagenomic pool of alkaline proteases JF - Microbial biotechnology Y1 - 2011 SN - 1432-0614 (E-Journal); 0171-1741 (Print); 0175-7598 (Print); 0340-2118 (Print) VL - Vol. 4 IS - Iss. 6 SP - 767 EP - 776 PB - Springer CY - Berlin ER - TY - JOUR A1 - Ribitsch, D. A1 - Heumann, S. A1 - Karl, W. A1 - Gerlach, J. A1 - Leber, R. A1 - Birner-Gruenberger, R. A1 - Gruber, K. A1 - Eiteljoerg, I. A1 - Remler, P. A1 - Siegert, Petra A1 - Lange, J. A1 - Maurer, Karl-Heinz A1 - Berg, G. A1 - Guebitz, G. M. A1 - Schwab, H. T1 - Extracellular serine proteases from Stenotrophomonas maltophilia: Screening, isolation and heterologous expression in E. coli JF - Journal of biotechnology N2 - A large strain collection comprising antagonistic bacteria was screened for novel detergent proteases. Several strains displayed protease activity on agar plates containing skim milk but were inactive in liquid media. Encapsulation of cells in alginate beads induced protease production. Stenotrophomonas maltophilia emerged as best performer under washing conditions. For identification of wash-active proteases, four extracellular serine proteases called StmPr1, StmPr2, StmPr3 and StmPr4 were cloned. StmPr2 and StmPr4 were sufficiently overexpressed in E. coli. Expression of StmPr1 and StmPr3 resulted in unprocessed, insoluble protein. Truncation of most of the C-terminal domain which has been identified by enzyme modeling succeeded in expression of soluble, active StmPr1 but failed in case of StmPr3. From laundry application tests StmPr2 turned out to be a highly wash-active protease at 45 °C. Specific activity of StmPr2 determined with suc-l-Ala-l-Ala-l-Pro-l-Phe-p-nitroanilide as the substrate was 17 ± 2 U/mg. In addition we determined the kinetic parameters and cleavage preferences of protease StmPr2. KW - Alginate beads KW - Stenotrophomonas maltophilia KW - Detergent protease Y1 - 2012 U6 - http://dx.doi.org/10.1016/j.jbiotec.2011.09.025 SN - 1873-4863 (E-Journal); 0168-1656 (Print) VL - 157 IS - 1 SP - 140 EP - 147 PB - Elsevier CY - Amsterdam ER - TY - PAT A1 - Maurer, Karl-Heinz A1 - O'Connell, Timothy A1 - Siegert, Petra A1 - Weber, Thomas A1 - Tondera, Susanne A1 - Hellmuth, Hendrik T1 - Flüssige Tensidzubereitung enthaltend Lipase und Phosphonat [Offenlegungsschrift] T1 - Liquid surfactant preparation containing lipase and phosphonate [Europäische Patentanmeldung / Internationale Patentanmeldung] Y1 - 2012 SP - 1 EP - 22 PB - Deutsches Patent- und Markenamt / Europäisches Patentamt / WIPO CY - München / Den Hague / Genf ER - TY - JOUR A1 - Martinez, Ronny A1 - Jakob, Felix A1 - Tu, Ran A1 - Siegert, Petra A1 - Maurer, Karl-Heinz A1 - Schwaneberg, Ulrich T1 - Increasing activity and thermal resistance of Bacillus gibsonii alkaline protease (BgAP) by directed evolution JF - Biotechnology and bioengineering Y1 - 2013 SN - 1097-0290 (E-Journal); 0006-3592 (Print); 0368-1467 (Print) VL - Vol. 110 IS - Iss. 3 SP - 711 EP - 720 PB - Wiley CY - Weinheim ER - TY - PAT A1 - Wieland, Susanne A1 - Siegert, Petra A1 - Spitz, Astrid A1 - Maurer, Karl-Heinz A1 - O'Connell, Timothy A1 - Prüser, Inken A1 - Schiedel, Marc-Steffen A1 - Eiting, Thomas A1 - Sendor-Müller, Dorota A1 - Bastigkeit, Thorsten A1 - Benda, Konstantin A1 - Müller, Sven T1 - Lagerstabiles flüssiges Wasch- oder Reinigungsmittel enthaltend Proteasen [Offenlegungsschrift] T1 - Storage-stable liquid washing or cleaning agent containing proteases [Europäische Patentanmeldung / Internationale Patentanmeldung] Y1 - 2011 SP - 1 EP - 25 PB - Deutsches Patent- und Markenamt / Europäisches Patentamt / WIPO CY - München / Den Hague / Genf ER - TY - PAT A1 - Bessler, Cornelius A1 - Evers, Stefan A1 - Maurer, Karl-Heinz A1 - Merkel, Marion A1 - Siegert, Petra A1 - Weber, Angrit A1 - Wieland, Susanne T1 - Leistungsverbesserte Proteasen und Wasch- und Reinigungsmittel enthaltend diese Proteasen [Offenlegungsschrift] T1 - Improved-performance proteases and detergents and cleaning agents comprising said proteases [Internationale Patentanmeldung] Y1 - 2009 SP - 1 EP - 41 PB - Deutsches Patent- und Markenamt / WIPO CY - München / Genf ER - TY - PAT A1 - Siegert, Petra A1 - Schwaneberg, Ulrich A1 - Martinez Moya, Ronny A1 - Merkel, Marion A1 - Spitz, Astrid A1 - Wieland, Susanne A1 - Hellmuth, Hendrik A1 - Maurer, Karl-Heinz T1 - Leistungsverbesserte Proteasevariante [Offenlegungsschrift] T1 - Performance-enhanced protease variant [Europäische Patentanmeldung / Internationale Patentanmeldung] Y1 - 2012 SP - 1 EP - 29 PB - Deutsches Patent- und Markenamt / Europäisches Patentamt / WIPO CY - München / Den Hague / Genf ER - TY - PAT A1 - O'Connell, Timothy A1 - Siegert, Petra A1 - Maurer, Karl-Heinz A1 - Schiedel, Marc-Steffen A1 - Vockenroth, Inga Kerstin T1 - Method for improving the cleaning action of a detergent or cleaning agent [Internationale Patentanmeldung] T1 - Verfahren zur Verbesserung der Reinigungsleistung eines Wasch- oder Reinigungsmittels Y1 - 2010 SP - 1 EP - 15 PB - WIPO CY - Genf ER -