caa a22 4500 60550119X CHVBK 20210128100559.0 cr unu---uuuuu 210128e20151101xx s 000 0 eng 10.1007/s00253-015-6758-z doi (NATIONALLICENCE)springer-10.1007/s00253-015-6758-z Characterization and mechanism insight of accelerated catalytic promiscuity of Sulfolobus tokodaii (ST0779) peptidase for aldol addition reaction [Elektronische Daten] [Rong Li, Bianca Perez, Hui Jian, Mads Jensen, Renjun Gao, Mingdong Dong, Marianne Glasius, Zheng Guo] A novel peptidase from thermophilic archaea Sulfolobus tokodaii (ST0779) is examined for its catalytic promiscuity of aldol addition, which shows comparable activity as porcine pancreatic lipase (PPL, one of the best enzymes identified for biocatalytic aldol addition) at 30°C but much accelerated activity at elevated temperature. The molecular catalytic efficiency kcat/Km (M−1s−1) of this thermostable enzyme at 55°C adds up to 140 times higher than that of PPL at its optimum temperature 37°C. The fluorescence quenching analysis depicts that the binding constants of PPL are significantly higher than those of ST0779, and their numbers of binding sites show opposite temperature dependency. Thermodynamic parameters estimated by fluorescence quenching analysis unveil distinctly different substrate-binding modes between PPL and ST0779: the governing binding interaction between PPL and substrates is hydrophobic force, while the dominating substrate-binding forces for ST0779 are van der Waals and H-bonds interactions. A reasonable mechanism for ST0779-catalyzed aldol reaction is proposed based on kinetic study, spectroscopic analysis, and molecular stereostructure simulation. This work represents a successful example to identify a new enzyme for catalytic promiscuity, which demonstrates a huge potential to discover and exploit novel biocatalyst from thermophile microorganism sources. Springer-Verlag Berlin Heidelberg, 2015 Aldol reaction nationallicence Sulfolobus tokodaii peptidase (ST0779) nationallicence Hydrolase nationallicence Catalytic promiscuity nationallicence Biocatalysis nationallicence Fluorescence spectroscopy nationallicence Li Rong Key Laboratory for Molecular Enzymology and Engineering, The Ministry of Education, Jilin University, 130021, Changchun, China aut Perez Bianca Department of Engineering, Faculty of Science and Technology, Aarhus University, Gustav Wied Vej 10, 8000, Aarhus, Denmark aut Jian Hui Key Laboratory for Molecular Enzymology and Engineering, The Ministry of Education, Jilin University, 130021, Changchun, China aut Jensen Mads Department of Chemistry and iNANO, Aarhus University, Langelandsgade 140, 8000, Aarhus C, Denmark aut Gao Renjun Key Laboratory for Molecular Enzymology and Engineering, The Ministry of Education, Jilin University, 130021, Changchun, China aut Dong Mingdong Interdisciplinary Nanoscience Center, Aarhus University, Gustav Wieds Vej 14, 8000, Aarhus C, Denmark aut Glasius Marianne Department of Chemistry and iNANO, Aarhus University, Langelandsgade 140, 8000, Aarhus C, Denmark aut Guo Zheng Department of Engineering, Faculty of Science and Technology, Aarhus University, Gustav Wied Vej 10, 8000, Aarhus, Denmark aut Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/22(2015-11-01), 9625-9634 0175-7598 99:22<9625 2015 99 253 https://doi.org/10.1007/s00253-015-6758-z text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00253-015-6758-z text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Li Rong Key Laboratory for Molecular Enzymology and Engineering, The Ministry of Education, Jilin University, 130021, Changchun, China aut NATIONALLICENCE 700 1- Perez Bianca Department of Engineering, Faculty of Science and Technology, Aarhus University, Gustav Wied Vej 10, 8000, Aarhus, Denmark aut NATIONALLICENCE 700 1- Jian Hui Key Laboratory for Molecular Enzymology and Engineering, The Ministry of Education, Jilin University, 130021, Changchun, China aut NATIONALLICENCE 700 1- Jensen Mads Department of Chemistry and iNANO, Aarhus University, Langelandsgade 140, 8000, Aarhus C, Denmark aut NATIONALLICENCE 700 1- Gao Renjun Key Laboratory for Molecular Enzymology and Engineering, The Ministry of Education, Jilin University, 130021, Changchun, China aut NATIONALLICENCE 700 1- Dong Mingdong Interdisciplinary Nanoscience Center, Aarhus University, Gustav Wieds Vej 14, 8000, Aarhus C, Denmark aut NATIONALLICENCE 700 1- Glasius Marianne Department of Chemistry and iNANO, Aarhus University, Langelandsgade 140, 8000, Aarhus C, Denmark aut NATIONALLICENCE 700 1- Guo Zheng Department of Engineering, Faculty of Science and Technology, Aarhus University, Gustav Wied Vej 10, 8000, Aarhus, Denmark aut NATIONALLICENCE 773 0- Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/22(2015-11-01), 9625-9634 0175-7598 99:22<9625 2015 99 253