Program members

Dr. Theodore G. Sotiroudis, Researcher A (Professor level) - Program Director

Dr. Georgios Skretas, Researcher D (Lecturer level)

Dr. Maria Zoumpanioti, Functional Research Scientist C

Research objectives

Enzyme Biotechnology constitutes the exploitation of enzyme biocatalysts either as living cells, cell extracts, or purified protein molecules for generating a useful product or service in a variety of industrial processes. Synthetic Biotechnology aims at engineering new cell functions and manipulating cellular processes for developing industrial production strains using a variety of tools, including proteomics, metabolomics, cell analytics, imaging, systems biology and bioinformatics, among others. These approaches open up exciting new avenues for innovation.

Recent emphasis of Enzyme Biotechnology activities in our Program is given on the study of oxidative enzymes (polyphenol oxidases, peroxidases, lipoxygenases) and antioxidant biomolecules in plant, food and microbial extracts. In this regard, we would like to understand the effect of critical oxidation processes on the quality and function of natural foods, such as olives, olive oil and the microalga Spirulina (Arthrospira), as well as to produce new products for the food, cosmetics and biomedical sectors. Accordingly, plant, food and microbial molecular components are extracted and examined for oxidative enzyme- and antioxidant-activities, as well as for enzyme inhibitory action. The formation of specific free radicals during enzyme and chemical oxidations, both in conventional and microemulsion systems, and the presence of free radical scavengers in cellular systems and food products is also presently studied using spin-trapping Electron Paramagnetic Resonance (EPR) spectroscopy. Enzymic transesterification of microalgae- and plant seed-oils is also being investigated for biodiesel production, as it produces high purity product and enables easy separation from the by-product, glycerol.

The goal of the Synthetic Biotechnology component of our Program is the development of engineered microbial cells with the ability to perform novel and complex functions by employing principles of Synthetic Biology. The Program utilizes simple organisms, such as the bacterium Escherichia coli and the microalga Spirulina platensis as "biological chasses" and seeks to evolve them into efficient "cell factories" for the production of high-value biotechnological products, and for the performance of industrially important processes, such as drug sensing and discovery. Genetic engineering techniques are applied in order to redesign and rearrange the genome of the organism of interest, while protein engineering approaches (directed protein evolution) are utilized so as to introduce novel functions in the cell. A key aspect of the work that is carried out is the design and development of high-throughput screening systems which are used to isolate the rare biomolecules and microbial strains that execute the desired function among large combinatorial libraries comprising millions of variants.

Indicative research projects currently underway include:     

• Understanding the action of oxidative processes on the quality of virgin olive oil and characterizing antioxidants in food products of the Mediterranean Diet.
• Development of new methodologies for the mass culture of microalgae (especially Spirulina) in sea-water. Production of new biochemicals for use in functional foods, cosmetics and biomedical applications, based on enzyme modifications of Spirulina bioactive constituents (phycocyanin, phycocyanobilin, sulphated polysaccharides), in collaboration with the Greek company ALGAE A.C. (Nigrita, Serres) (http://www.spirulina.gr).
  
• Enzymic production of biodiesel from microalgae and greek halophyte seed oils.
  
• Directed evolution of small-molecule therapeutics against protein misfolding diseases, such as neurodegenerative disorders (Alzheimer's and Parkinson's disease), type II diabetes, and cancer.
  
• Development of enzyme-based molecular sensors of protein conformation and dynamics.
  
• Screening of metagenomic libraries for the identification of thermostable enzymes for industrial processes.
  
• Screening of libraries of natural products for the discovery of compounds with anti-angiogenic effects.
  
• Engineering of E. coli for enhanced production of hard-to-express recombinant proteins.
  
• Engineering of Spirulina platensis for high-level production of high-added value products.

Representative  publications:

1.   Tzika, E., Christoforou, M., Pispas, S., Zervou,M., Papadimitriou, V., Sotiroudis, T.G., Leontidis, E. and Xenakis, A. (2011) Influence of nanoreactor environment and substrate location on the activity of horseradish peroxidase in olive oil-based w/o microemulsions.  Langmuir, 27, 2692-2700.

2.   Papadimitriou, V., Tzika, E. D.,   Pispas,  S.,  Sotiroudis, T. G. and   Xenakis, A., (2011) Microemulsions based on virgin olive oil: a model biomimetic system for studying native oxidative enzymatic activities. Colloids and Surfaces A: Physicochemical and Engineering Aspects, in press.

3. Makino, T., Skretas, G., Kang, T.H., and Georgiou, G. (2010) Comprehensive Engineering of Escherichia coli for Enhanced Expression of IgG antibodies. Metabolic Engineering, Dec 3. [Epub ahead of print].

4.   Xenakis, A., Papadimitriou, V., and Sotiroudis, T.G. (2010) Colloidal structures in natural oils. Curr.Opin.Colloid Interface Sci. 15, 55-60.

5. Zoumpanioti M, Merianou E, Karandreas T, Stamatis H, Xenakis A. (2010) Esterification of phenolic acids catalyzed by lipases immobilized in organogels. Biotechnol. Lett. 32(10):1457-62

6.   Sotiroudis, V.T., Sotiroudis, T.G. and Kolisis, F.N. (2010) The potential of biodiesel production from fatty acid methyl esters of some European/ Mediterranean and Cosmopolitan halophyte seed oils.  Journal of ASTM International, 7 (3) 1-9

7. Skretas, G., and Georgiou., G. (2009) Genetic analysis of G protein-coupled receptor expression in Escherichia coli: Inhibitory role of DnaJ on the membrane integration of the human central cannabinoid receptor. Biotechnology and Bioengineering. 102(2) : 357-367

8   Sotiroudis, T.G. and Kyrtopoulos, S.A. (2008) Anticarcinogenic compounds of olive oil and related biomarkers. Eur.J.Nutr. 47(Suppl 2) 69-72.

9.   Skretas, G., and Georgiou, G. (2008) Engineering G protein-coupled receptor expression in bacteria. Proc. Natl. Acad. Sci. USA. 105(39), 14747-14748

10. Papadimitriou V, Pispas S, Syriou S, Pournara A, Zoumpanioti M, Sotiroudis TG, Xenakis A. (2008) Biocompatible microemulsions based on limonene: formulation, structure, and applications. Langmuir, 24(7):3380-6

11. Papadimitriou, V., Sotiroudis, T.G., Xenakis, A. (2007) Olive oil microemulsions: enzymatic activities and structural characteristics. Langmuir, 23, 2071-2077.

12. Skretas, G., Meligova, A., Villalonga-Barber, C., Mitsiou, D.J., Alexis, M.N., Micha-Screttas, M., Steele, B.R., Screttas, C.G., and Wood, D.W. (2007) Engineered chimeric enzymes as facile tools for pharmaceutical discovery: Construction of simple bacterial screens for the detection, discovery and assessment of estrogen receptor modulators. J. Amer. Chem. Soc., 129,  8443-8457.

13. Papadimitriou, V., Sotiroudis, T.G., and Xenakis, A. (2005) Olive oil microemulsions as a biomimetic medium for enzymatic studies: oxidation of oleuropein. J.Amer.Oil Chem.Soc. 82, 335-340.

14. Skretas, G., and Wood, D.W. (2005) Regulation of protein activity with small-molecule-controlled inteins. Protein Science. 14: 523-532.

15. Karapitta, D.D, Sotiroudis,T.G, Papadimitriou,.A and, Xenakis, A. (2001) A homogeneous enzyme immunoassay for triiodothyronine in serum. Clin. Chem. 47, 569-57.

Issued patents:

1. Karapitta, C., Sotiroudis, T.G., Xenakis, A. (Inventors) (2003) Development of a new homogeneous immunoenzymic method for the production of a clinical laboratory test system (kit) for the quantification of thyroxine and triiodothyronine in human serum, utilizing polyiodothyronine conjugated with glycogen phosphorylase b. Hellenic Industrial Property Organisation, Patent 20000100255/1004119 (with participation of MEDICON HELLAS SA and NHRF).

2. Wood, D.W. and Skretas, G. (2009) Bacterial ligand-binding sensor. United States Patent 7592144.

Current funding sources-Participation in research projects

2011-2015. FP7 Cooperation KBBE - Food, Agriculture and Fisheries, Biotechnology General Call for Proposals. "Systematic screening for novel hydrolases from hot environments", Scientific Coordinator:  Prof. Xu Peng.

2011-2015. FP7-PEOPLE-RG. Marie Curie Actions - International Reintegration Grants. "Directed evolution of small-molecule cancer therapeutics", Scientific Coordinator: Dr. Georgios Skretas.   

2011-2015. National Program "Cooperation" "Development of novel angiogenesis-modulating pharmaceuticals by screening of natural compounds and synthetic analogues", Scientific Coordinator: Prof. A-L Skaltsounis.

2010-2013. FP7-REGPOT-2009-2 "Reinforcement of the Faculty of Chemistry, University of Belgrade, towards becoming a Center of Excellence in the region of WB for Molecular Biotechnology and Food research (FCUB-ERA)" Scientific Coordinator: Prof. Tanja Cirkovic Velickovic.