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Rando Duvikene

Institution(s) and Institute/Institutes  Tallinn University, Institute of Natural and Health Sciences

Head of the research team:  Prof. Rando Tuvikene

Members (including doctoral students): Marju Robal, Renu Geetha Bai; PhD students: Kadri Saluri, Musa Howlader, Amal Dharmapriya Premarathna Deliwala Ambegoda Gedara, Sanjida Hymayun.

Key words characterizing the research group's activities: seaweeds, polysaccharides, pigments, plant active substances, marine bacteria, biotechnology, enzyme technology, rheology, molecular structure of biopolymers, functional materials, biological activity, production processes and technologies.

Activity overview

The research group deals with the isolation of biopolymers and low-molecular-weight active substances from marine biomass and the purification, functionalization and clarification of the properties of these substances. The goal is to maximize the use of natural resources and thereby reduce the amount of industrial waste. Novel functional materials and visco-elastic systems are being developed on the basis of components originating from marine biomass. The research group has particularly extensive experience in the field of saccharides (especially galactans, fucans, alginates, chitosans), has been engaged in the valorization of seaweed (red, brown and green algae), mussels, shrimps, sea cucumbers, etc., including applying enzyme technologies and other environmentally friendly procedures.

Total research group funding 2020 (to the nearest 0.1 MEUR): 0.3 MEUR.

Business cooperation partners and topics in the last 5 years

• Est-Agar AS (Estonia, 2021 a strategic cooperation agreement was also signed) – furcellaran production technology developments and quality control of production products.
• SINTEF (Norway) - Novel applications of seaweed-derived components.
• Vetik OÜ (Estonia) - technology developments for the separation and purification of pigment substances originating from algae.
• Furcella OÜ (Estonia) – use of functional biomolecules from algae in cosmetic products.
• Chemi-Pharm AS (Estonia) - the use of saccharides in deodorants.

Key equipment, know-how, protected intellectual property

• Specialized equipment for crushing marine biomass (cooled mills, ball mills, including cryogenic grinding at liquid nitrogen temperature), for pretreatment (including the possibility of high-intensity sonication) and fractional extraction of the components contained in them.
• Dynamic rheometer (geometries: PP, CP, concentric cylinders) - detection of rheological properties of viscoelastic materials/systems (including in temperature gradient), detection of melting and gelation of gels.
• HPLC-RI (SEC) - determination of molecular weights (Mp, Mw, Mn, Mz) and polydispersity of polysaccharides, quantification of polysaccharides.
• HPLC-PDA (SEC) - quantification of active substances, determination of molecular weights of proteins.
• HPLC-FL (fluorescence detector) - quantification of pigments.
• HPLC-PAD (electrochemical detector in pulse amperometry mode) - quantification of low molecular saccharides in pM concentration zones, quantification of oligosaccharides, detection of the monomer composition of polysaccharides after hydrolysis.
• HPLC-ELSD - Quantification of biopolymers.
• GC-MS/FID - Determination of Monomer Composition of Polysaccharides.
• Automated ultrafiltration system (sensors: pH, conductivity, UV absorbance, flow rates, pressures, retentate and ultrafiltrate masses) - large-scale fractionation and purification of extracts.
• FT-IR/FT-Raman (1064 nm laser) - characterization of raw materials, biosamples and industrial products, quality control, determination of the degree of acetylation of chitin and chitosan.
• Nephelometry, Peltier thermostated spectrophotometry - evaluation of the turbidity of beverages, detection of clarification efficiency of preparations, optical phenomena accompanying phase transitions in gel-salt systems.
• Evaluation of bioeffects on eukaryotic cell cultures, enzymatic and chemical modification of polysaccharides, detection of tyrosinase inhibition (skin bleaching properties), evaluation of anticoagulant activity (aPTT, TT), antioxidant activity, antibacterial activity.

The device models are visible at: https://www.tlu.ee/lti/teaduslaborid/lti-molekulaarteaduste-labori-oluline-teadusaparatuur

Services offered

• Extraction of biomolecules from biomass.
• Functionalization of biopolymers.
• Comprehensive characterization of polysaccharides.

Desires for cooperation with Estonian companies

• Valorization of marine biomass, including development of necessary technological solutions.
• Topics related to the isolation, purification, functionalization and properties of polysaccharides.
• Development of functional viscoelastic systems based on polysaccharides.

Priit Väljamäe

Head of the research group: associate professor Priit Väljamäe
Contact details: Riga 23b – 202, Tel: 737 5823, priit.valjamae@ut.ee

Research directions
• Enzymatic degradation of recalcitrant polysaccharides (cellulose and chitin).
• Redox reactions during the degradation of lignocellulose

Description of the research

People
Priit Väljamäe, associate professor, PhD, priit.valjamae@ut.ee
Silja Kuusk, researcher, PhD, silja.kuusk@ut.ee
Jürgen Jalak, researcher, PhD, jyrgen.jalak@ut.ee
Alexander Rannar, Master's student
Alexey Nesterovich, undergraduate student
Alisa Kamnerov, undergraduate student

Research work

• Structural polysaccharides - cellulose and chitin - contain a huge reserve of renewable carbon. Their enzyme-mediated beneficiation offers a green and sustainable alternative to the traditional oil-based industry. Unfortunately, the crystalline structure of cellulose and chitin makes them difficult for enzymes to break down, hence the collective name recalcitrant polysaccharides. The development of optimal enzyme mixtures requires a thorough knowledge of the action mechanisms of individual enzymes. The detailed description of the action mechanism and kinetics of enzymes is the main direction of our research.
• Enzymatic degradation of recalcitrant polysaccharides (cellulose and chitin).

Due to the crystalline structure of cellulose and chitin, their enzymatic degradation is a heterogeneous catalysis at the solid-liquid interface, which includes a series of different intermediate steps. An important step towards increasing the speed of the overall reaction is to find out the rate-limiting intermediate step (the so-called bottleneck). Our research group's so-called the strength is the development and implementation of novel methods allowing the measurement of the rate of individual intermediate steps. The above approach has enabled us to determine the processivity of glycoside hydrolases (different cellulases and chitinases) and the values of constants characterizing the rate of various intermediate steps such as association, dissociation and glycosidic bond hydrolysis. In addition to traditional glycoside hydrolases, we have also focused on lytic polysaccharide monooxygenases (LPMOs). LPMOs are recently discovered redox enzymes involved in the degradation of recalcitrant polysaccharides. Understanding their operation and kinetics holds great potential for increasing the efficiency of degradation and modification of recalcitrant polysaccharides.

• Redox reactions in lignocellulose degradation

LPMOs need both electrons (reduction) and H2O2/O2 co-substrate (oxidation) to function. The recognition of the importance of LPMOs has drawn increased attention to the involvement of redox reactions in the degradation of lignocellulose (the main component of the plant cell envelope). Here, we focus on the development of enzyme cascades that would enable optimal conditions for the activity and stability of LPMOs in a complex redox-active environment. In addition to LPMOs, we are interested in enzymes active on lignin and its degradation products, such as various laccases, peroxidases and oxidases.

A selection of publications

1. Kont, R., Bissaro, B., Eijsink, VGH, and Väljamäe, P. (2020) Kinetic insights into the peroxygenase activity of cellulose-active lytic polysaccharide monooxygenases (LPMOs). Nat. Commun. 11:5786
2. Vermaas, JV, Kont, R., Beckham, GT, Crowley, MF, Gudmundsson, M., Sandgren, M., Ståhlberg, J., Väljamäe, P., and Knott, BC (2019) The dissociation mechanism of processive cellulase. Proc. Natl. Acad. Sci. USA. 116, 23061-23067
3. Kont, R., Pihlajaniemi, V., Borisova, AS, Aro, N., Marjamaa, K., Loogen, J., Büchs, J., Eijsink, VGH, Kruus, K., and Väljamäe, P. (2019) The liquid fraction from hydrothermal pretreatment of wheat straw provides lytic polysaccharide monooxygenases with both electrons and H2O2 co-substrate. Biotechnol. Biofuels 12:235
4. Kuusk, S., Kont, R., Kuusk, P., Heering, A., Sørlie, M., Bissaro, B., Eijsink, VGH, and Väljamäe, P. (2019) Kinetic insights into the role of the reductant in H2O2-driven degradation of chitin by a bacterial lytic polysaccharide monooxygenase. J. Biol. Chem. 294, 1516-1528
5. Kuusk, S., Bissaro, B., Kuusk, P., Forsberg, Z., Eijsink, VGH, Sørlie, M., and Väljamäe, P. (2018) Kinetics of H2O2-driven degradation of chitin by a bacterial lytic polysaccharide monooxygenase. J. Biol. Chem. 293, 523-531
6. Kuusk, S., and Väljamäe, P. (2017) When substrate inhibits and inhibitor activates: implications of β-glucosidases. Biotechnol. Biofuels 10:7
7. Kurašin, M., Kuusk, S., Kuusk, P., Sørlie, M., and Väljamäe, P. (2015) Slow off-rates and strong product binding are required for processivity and efficient degradation of recalcitrant chitin by family 18 in chitinas. J. Biol. Chem. 290, 29074-29085
8. Kuusk, S., Sørlie, M., and Väljamäe, P. (2015) The predominant molecular state of bound enzyme determines the strength and type of product inhibition in the hydrolysis of recalcitrant polysaccharides by processive enzymes. J. Biol. Chem. 290, 11678-11691
9. Jalak, J., Kurašin, M., Teugjas, H., Väljamäe, P. (2012) Endo-exo synergism in cellulose hydrolysis revisited. J. Biol. Chem. 287, 28802-28815.
10. Kurašin, M., Väljamäe, P. (2011) Processivity of cellobiohydrolases is limited by the substrate. J. Biol. Chem. 286, 169–177.

More publications.

Indrek Rei

Group leader: Dr. Indrek Rei

The Laboratory for the Analysis of Complex Mixtures is a group of researchers at KBFI that focuses on the analysis of multicomponent samples using nuclear magnetic resonance spectroscopy (NMR/NMR). The group consists of 8 people, 4 of whom have PhDs. KSA operates in two research directions: development of NMR methodology towards greater sensitivity; and chemical analysis of complex multicomponent samples and macromolecules. Examples of ongoing projects:

• PSG11 "Quantitative analysis of cancer markers in urine by hyperpolarized NMR" - development of a methodology for the discovery of new cancer markers in urine samples. Hyper-polarization helps to see analytes below the detection limit of conventional NMR analysis (ETIS)
• COVSG7 "Saccharides with antiviral effect: applications in deodorants, cosmetic products and pharmaceutical formulations" - In cooperation with Tallinn University, we apply modern NMR techniques to understand the structure of saccharides with antiviral effect found in Estonian nature and to support the development of products with virus-inhibiting properties. (ETIS)

KBFI-KSA carries out commissioned work on NMR analysis for companies on a daily basis and conducts research cooperation with many research institutions and research groups in Estonia and abroad. In the field of wood chemistry, in cooperation with TalTech, we have developed competence for NMR analysis of lignin preparations, supporting the work of Estonian scientists and companies in the valuation of wood components.