We are using efficient expression systems to produce heterologous proteins from pro- and eukaryotic organisms in Escherichia coli. We have expressed glucoside hydrolases (e.g. levansucrases, α-glucosidases) for their purification, characterization and enzymatic synthesis of saccharides. We have studied bacterial and yeast enzymes that can be applied to synthesize novel potential prebiotics and functional saccharides from table sugar (sucrose). We are using levansucrase from a bacterium Pseudomonas syringae to synthesize polymeric levan and endo-levanase of Bacteroides thetaiotaomicron to cleave it into short-chain oligosaccharides. These saccharides stimulate the growth of beneficial gut bacteria. We have solved the 3D structure of the maltase from a yeast Blastobotys adeninivorans and discovered its ability to synthesize various rare bioactive oligosaccharides. Immobilization of enzymes to solid support enables their repeated use. Therefore, with the same amount of enzyme it is possible to obtain higher yields. We are investigating the properties and stability of biotechnologically relevant lectin-tagged enzymes.
Header image: The structure of the maltase BaAG2 of yeast Blastobotrys adeninivorans with acarbose in the reaction centre. Author of the image: Karin Ernits
Pereira, R., Ugarte-Berzal, E., Vandooren, J., Nylander, K., Martens, E., Van Mellaert, L., Van Damme, J., Vranckx, J. J., Matthys, P., Alamäe, T., Phillipson, M., Visnapuu, T., & Opdenakker, G. (2022). Chlorite-Oxidized Oxyamylose (COAM) Has Antibacterial Activity and Positively Affects Skin Wound Healing. Journal of inflammation research, 15, 4995–5008. DOI: 10.2147/JIR.S375487
Lust Rauno, Nerut Jaak, Gadegaonkar Sharvari S., Kasak Kuno, Espenberg Mikk, Visnapuu Triinu, Mander Ülo (2022) Single-chamber microbial electrosynthesis reactor for nitrate reduction from waters with a low-electron donors’ concentration: from design and set-up to the optimal operating potential. Frontiers in Environmental Science, 10, DOI: 10.3389/fenvs.2022.938631
Young, Ian D.; Nepogodiev, Sergey A.; Black, Ian M.; Le Gall, Gwenaelle; Wittmann, Alexandra; Latousakis, Dimitrios; Visnapuu, Triinu; Azadi, Parastoo; Field, Robert A.; Juge, Nathalie; Kawasaki, Norihito (2022). Lipopolysaccharide associated with β-2,6 fructan mediates TLR4-dependent immunomodulatory activity in vitro. Carbohydrate Polymers, 118606. DOI: 10.1016/j.carbpol.2021.118606
Ernits, Karin; Kjeldsen, Christian; Persson, Karina; Grigor, Eliis; Alamäe, Tiina; Visnapuu, Triinu (2021). Structural Insight into a Yeast Maltase—The BaAG2 from Blastobotrys adeninivorans with Transglycosylating Activity. Journal of Fungi, 7 (10), 816. DOI: 10.3390/jof7100816
Visnapuu, Triinu; Meldre, Aivar; Põšnograjeva, Kristina; Viigand, Katrin; Ernits, Karin; Alamäe, Tiina (2020). Characterization of a Maltase from an Early-Diverged Non-Conventional Yeast Blastobotrys adeninivorans. International Journal of Molecular Sciences, 21 (1), 297. DOI: 10.3390/ijms21010297
Visnapuu, Triinu; Teze, David; Kjeldsen, Christian; Lie, Aleksander; Duus, Jens Ollgaard; Andre-Miral, Corinne; Pedersen, Lars Haastrup; Stougaard, Peter; Svensson, Birte (2020). Identification and Characterization of a β-N-Acetylhexosaminidase with a Biosynthetic Activity from the Marine Bacterium Paraglaciecola hydrolytica S66T. International Journal of Molecular Sciences, 21 (2), 417. DOI: 10.3390/ijms21020417
Ernits, Karin; Eek, Priit; Lukk, Tiit; Visnapuu, Triinu; Alamäe, Tiina (2019). First crystal structure of an endo-levanase – the BT1760 from a human gut commensal Bacteroides thetaiotaomicron. Scientific Reports, 9 (1), 8443. DOI: 10.1038/s41598-019-44785-0
Short communication of the research topic in English: https://researchinestonia.eu/2019/09/26/what-should-i-eat-to-make-my-gut-bacteria-happy/
The following student research projects were completed in the research group, containing summary in English:
https://akadeemiake.ee/2021/06/polusahhariidse-levaani-moju-hindamine-valitud-bakteriliikide-kasvule-ning-tekkinud-produktide-analuus/ In 2020 the paper received a special prize at the National Contest for Student Researchers (organized by the Estonian Research Council).
https://akadeemiake.ee/2021/10/ebatraditsioonilise-parmi-blastobotrys-adeninivorans-maltaasi-ag2-aktiivsuse-maaramine-ja-biokeemiliste-omaduste-iseloomustamine/ In 2021 the paper received the 1st prize in the high school level at the National Contest for Student Researchers (organized by the Estonian Research Council) and a special prize at The European Union Contest for Young Scientists.
Characterization of the activities and biochemical parameters of maltase AG2 from the non-conventional yeast Blastobotrys adeninivorans, short video in English (YouTube)
Characterization of the activities and biochemical parameters of maltase AG2 from the non-conventional yeast Blastobotrys adeninivorans, in Estonian (YouTube)