TY - JOUR A1 - Tran, Quang Hon A1 - Bongaerts, Johannes A1 - Vlad, Dorina A1 - Unden, Gottfried T1 - Requirement for the proton-pumping NADH dehydrogenase I of Escherichia coli in respiration of NADH to fumarate and its bioenergetic implications JF - European journal of biochemistry Y1 - 1997 SN - 0014-2956 VL - Vol. 244 IS - Iss. 1 SP - 155 EP - 160 ER - TY - JOUR A1 - Schmitz, M. A1 - Hirsch, E. A1 - Bongaerts, Johannes A1 - Takors, Ralf T1 - Pulse experiments as a prerequisite for the quantification of in vivo enzyme kinetics in aromatic amino acid pathway of Eschericia coli JF - Biotechnology progress Y1 - 2002 SN - 1520-6033 (E-Journal); 8756-7938 (Print) VL - Vol. 18 IS - Iss. 5 SP - 935 EP - 941 ER - TY - JOUR A1 - Gerigk, M. A1 - Bujnicki, R. A1 - Ganpo-Nkwenkwa, E. A1 - Bongaerts, Johannes A1 - Sprenger, G. A1 - Takors, Ralf T1 - Process control for enhanced L-phenylalanine production using different recombinant Escherichia coli strains JF - Biotechnology and bioengineering Y1 - 2002 SN - 1097-0290 (E-Journal); 0006-3592 (Print) VL - Vol. 80 IS - Iss. 7 SP - 746 EP - 754 ER - TY - JOUR A1 - Falkenberg, Fabian A1 - Bott, Michael A1 - Bongaerts, Johannes A1 - Siegert, Petra T1 - Phylogenetic survey of the subtilase family and a data-mining-based search for new subtilisins from Bacillaceae JF - Frontiers in Microbiology N2 - The subtilase family (S8), a member of the clan SB of serine proteases are ubiquitous in all kingdoms of life and fulfil different physiological functions. Subtilases are divided in several groups and especially subtilisins are of interest as they are used in various industrial sectors. Therefore, we searched for new subtilisin sequences of the family Bacillaceae using a data mining approach. The obtained 1,400 sequences were phylogenetically classified in the context of the subtilase family. This required an updated comprehensive overview of the different groups within this family. To fill this gap, we conducted a phylogenetic survey of the S8 family with characterised holotypes derived from the MEROPS database. The analysis revealed the presence of eight previously uncharacterised groups and 13 subgroups within the S8 family. The sequences that emerged from the data mining with the set filter parameters were mainly assigned to the subtilisin subgroups of true subtilisins, high-alkaline subtilisins, and phylogenetically intermediate subtilisins and represent an excellent source for new subtilisin candidates. Y1 - 2022 U6 - http://dx.doi.org/10.3389/fmicb.2022.1017978 SN - 1664-302X VL - 2022 IS - 13 PB - Frontiers CY - Lausanne ER - TY - JOUR A1 - Unden, G. A1 - Becker, S. A1 - Bongaerts, Johannes A1 - Schirawski, J. A1 - Six, S. T1 - Oxygen regulated gene expression in facultatively anaerobic bacteria JF - Antonie van Leeuwenhoek Y1 - 1994 SN - 0003-6072 (Print) ; 1572-9699 (online) VL - Vol. 66 IS - Iss. 1-3 SP - 3 EP - 22 ER - TY - JOUR A1 - Degering, Christian A1 - Eggert, Thorsten A1 - Puls, Michael A1 - Bongaerts, Johannes A1 - Evers, Stefan A1 - Maurer, Karl-Heinz A1 - Jaeger, Karl-Erich T1 - Optimization of protease secretion in Bacillus subtilis and Bacillus licheniformis by screening of homologous and herologous signal peptides JF - Applied and environmental microbiology N2 - Bacillus subtilis and Bacillus licheniformis are widely used for the large-scale industrial production of proteins. These strains can efficiently secrete proteins into the culture medium using the general secretion (Sec) pathway. A characteristic feature of all secreted proteins is their N-terminal signal peptides, which are recognized by the secretion machinery. Here, we have studied the production of an industrially important secreted protease, namely, subtilisin BPN′ from Bacillus amyloliquefaciens. One hundred seventy-three signal peptides originating from B. subtilis and 220 signal peptides from the B. licheniformis type strain were fused to this secretion target and expressed in B. subtilis, and the resulting library was analyzed by high-throughput screening for extracellular proteolytic activity. We have identified a number of signal peptides originating from both organisms which produced significantly increased yield of the secreted protease. Interestingly, we observed that levels of extracellular protease were improved not only in B. subtilis, which was used as the screening host, but also in two different B. licheniformis strains. To date, it is impossible to predict which signal peptide will result in better secretion and thus an improved yield of a given extracellular target protein. Our data show that screening a library consisting of homologous and heterologous signal peptides fused to a target protein can identify more-effective signal peptides, resulting in improved protein export not only in the original screening host but also in different production strains. Y1 - 2010 U6 - http://dx.doi.org/10.1128/AEM.01146-10 SN - 1098-5336 (E-Journal); 0003-6919 (Print); 0099-2240 (Print) VL - 76 IS - 19 SP - 6370 EP - 6378 PB - American Society for Microbiology CY - Washington, DC ER - TY - JOUR A1 - Unden, G. A1 - Becker, S. A1 - Bongaerts, Johannes A1 - Holighaus, G. A1 - Schirawski, J. A1 - Six, S. T1 - O2-sensing and O2-dependent gene regulation in facultatively anaerobic bacteria JF - Archives of microbiology Y1 - 1995 SN - 1432-072X (E-Journal); 0003-9276 (Print); 0302-8933 (Print) VL - Vol. 164 IS - Iss. 2 SP - 81 EP - 90 ER - TY - JOUR A1 - Haeger, Gerrit A1 - Probst, Johanna A1 - Jaeger, Karl-Erich A1 - Bongaerts, Johannes A1 - Siegert, Petra T1 - Novel aminoacylases from Streptomyces griseus DSM 40236 and their recombinant production in Streptomyces lividans JF - FEBS Open Bio N2 - Amino acid-based surfactants are valuable compounds for cosmetic formulations. The chemical synthesis of acyl-amino acids is conventionally performed by the Schotten-Baumann reaction using fatty acyl chlorides, but aminoacylases have also been investigated for use in biocatalytic synthesis with free fatty acids. Aminoacylases and their properties are diverse; they belong to different peptidase families and show differences in substrate specificity and biocatalytic potential. Bacterial aminoacylases capable of synthesis have been isolated from Burkholderia, Mycolicibacterium, and Streptomyces. Although several proteases and peptidases from S. griseus have been described, no aminoacylases from this species have been identified yet. In this study, we investigated two novel enzymes produced by S. griseus DSM 40236ᵀ . We identified and cloned the respective genes and recombinantly expressed an α-aminoacylase (EC 3.5.1.14), designated SgAA, and an ε-lysine acylase (EC 3.5.1.17), designated SgELA, in S. lividans TK23. The purified aminoacylase SgAA was biochemically characterized, focusing on its hydrolytic activity to determine temperature- and pH optima and stabilities. The aminoacylase could hydrolyze various acetyl-amino acids at the Nα -position with a broad specificity regarding the sidechain. Substrates with longer acyl chains, like lauroyl-amino acids, were hydrolyzed to a lesser extent. Purified aminoacylase SgELA specific for the hydrolysis of Nε -acetyl-L-lysine was unstable and lost its enzymatic activity upon storage for a longer period but could initially be characterized. The pH optimum of SgELA was pH 8.0. While synthesis of acyl-amino acids was not observed with SgELA, SgAA catalyzed the synthesis of lauroyl-methionine. KW - Streptomyces lividans KW - recombinant expression KW - Streptomyces griseus KW - ε-lysine acylase KW - α-aminoacylase Y1 - 2023 U6 - http://dx.doi.org/10.1002/2211-5463.13723 SN - 2211-5463 N1 - Corresponding author: Petra Siegert VL - 13 IS - 12 SP - 2224 EP - 2238 PB - Wiley CY - Hoboken, NJ ER - TY - JOUR A1 - Falkenberg, Fabian A1 - Voß, Leonie A1 - Bott, Michael A1 - Bongaerts, Johannes A1 - Siegert, Petra T1 - New robust subtilisins from halotolerant and halophilic Bacillaceae JF - Applied Microbiology and Biotechnology N2 - The aim of the present study was the characterisation of three true subtilisins and one phylogenetically intermediate subtilisin from halotolerant and halophilic microorganisms. Considering the currently growing enzyme market for efficient and novel biocatalysts, data mining is a promising source for novel, as yet uncharacterised enzymes, especially from halophilic or halotolerant Bacillaceae, which offer great potential to meet industrial needs. Both halophilic bacteria Pontibacillus marinus DSM 16465ᵀ and Alkalibacillus haloalkaliphilus DSM 5271ᵀ and both halotolerant bacteria Metabacillus indicus DSM 16189 and Litchfieldia alkalitelluris DSM 16976ᵀ served as a source for the four new subtilisins SPPM, SPAH, SPMI and SPLA. The protease genes were cloned and expressed in Bacillus subtilis DB104. Purification to apparent homogeneity was achieved by ethanol precipitation, desalting and ion-exchange chromatography. Enzyme activity could be observed between pH 5.0–12.0 with an optimum for SPPM, SPMI and SPLA around pH 9.0 and for SPAH at pH 10.0. The optimal temperature for SPMI and SPLA was 70 °C and for SPPM and SPAH 55 °C and 50 °C, respectively. All proteases showed high stability towards 5% (w/v) SDS and were active even at NaCl concentrations of 5 M. The four proteases demonstrate potential for future biotechnological applications. KW - Biotechnological application KW - Bacillaceae KW - Subtilisin KW - Subtilases KW - Halotolerant protease Y1 - 2023 U6 - http://dx.doi.org/10.1007/s00253-023-12553-w SN - 1432-0614 N1 - Corresponding author: Petra Siegert VL - 107 SP - 3939 EP - 3954 PB - Springer Nature CY - Berlin ER - TY - JOUR A1 - Wilming, Anja A1 - Begemann, Jens A1 - Kuhne, Stefan A1 - Regestein, Lars A1 - Bongaerts, Johannes A1 - Evers, Stefan A1 - Maurer, Karl-Heinz A1 - Büchs, Jochen T1 - Metabolic studies of γ-polyglutamic acid production in Bacillus licheniformis by small-scale continuous cultivations JF - Biochemical engineering journal Y1 - 2013 SN - 1873-295X (E-Journal); 1369-703X (Print) VL - Vol. 73 SP - 29 EP - 37 PB - Elsevier CY - Amsterdam ER -