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Research Group 3 - RG3 - dr Nenad Milosavić

Research description:

Pseudomonas cepacia lipaza Despite their many favorable qualities, the marginal stability of biocatalysts in many types of reaction media often has prevented or delayed their implementation for industrial-scale synthesis of fine chemicals and pharmaceuticals. Biocatalysts are inherently labile; therefore their operational stability is of paramount importance for any bioprocess. There is great interest in understanding effects of solution conditions on protein stability, as well as in developing strategies to improve protein stability in the desired reaction media. For biocatalytic processes to become more viable alternatives to chemical ones, new strategies for stabilizing enzymes in a variety of media must be developed. Chemical stabilization methods using additives, immobilization or a combination of both have all proven successful, although immobilization enables easier post-reaction separation of products and biocatalyst from reaction mixtures. Often these methods are developed on an individual basis for each biocatalyst process. Fundamental studies of the effects of solution conditions (e.g. temperature, pH and cosolute composition) on enzyme stability will allow guiding principles for stabilization strategies to be established and alleviate the need for case-by-case stabilization. Recent developments to modify enzymes for use in organic synthesis have targeted several areas. These include altering the reaction mechanism of the enzyme to catalyse new reactions, switching substrate specificity, expanding substrate specificity, and improving substrate specificity, such as enantioselectivity in kinetic resolutions.

This makes for substantial process energy savings and reduced manufacturing costs. Also, enzymes practically do not present disposal problems since, being mostly proteins and peptides, they are biodegradable and easily removed from contaminated streams. This unique set of advantageous features of enzymes as catalysts has been exploited since the 1960s and several enzyme-catalyzed processes have been successfully introduced to industry, e.g. in the production of certain foodstuffs, pharmaceuticals and agrochemicals, but now also increasingly to organic chemical synthesis. Enzymatic methods are now widely accepted as being as much a part of oligosaccharide synthesis as more traditional chemical methods.

Given the often long-winded protection regimes required for most chemical glycosylations, the use of enzyme-catalyzed one-step systems is clearly attractive. The development of stereoselective methods for the synthesis of glycosidic linkages presents a considerable challenge to synthetic chemists. Chemical syntheses of the glycosidic moieties are mainly based on time-consuming protection and deprotection strategies, activation or metal catalysis, but are often accompanied by formation of unwanted diastereomers and low yields. However, these difficulties can be overcome enzymatically.

Research skills and methods applied by the group:
Group members and ongoing projects:

Ratko Jankov, senior researcher
Projects: Enzymatic synthesis of physiological active glycosides,
Immobilization and chemical stabilization of hydrolases.

dr Ziyad Tantoush, part-time PhD student
Project: Novel phenolic mediators of enzymatic cross-linking

Aleksandra Dimitrijević, PhD student
Project: Production, isolation and stabilization of microbial lipases and their utilization in organic sythesis

Dušan Veličković, PhD student
Project: Glucosidase purification and enzymatic sythesis of physiological active glycosides

Selected recent publications:
  1. Associations of Invertors and Authors of Technical Improvements Awards Silver Medal with Nicola Tesla’s Face; Belgrade, No 089-10, May 2010.
  2. Milosavic N, Prodanovic R, Macroporous poly (GMA-co EGDMA) for Enzyme stabilization, book chapter, Springer Science copyright year 2011.
  3. Žuža, M., Milosavić, N., Knežević-Jugović, Z.; Immobilization of modified penicillin G acylase on Sepabeads carriers Chemical Papers 63 (2),117-124, 2009.
  4. Gavrovic-Jankulovic, M., Spasic, M., Velickovic, T. C., Stojanovic, M., Inic-Kanada, A., Dimitrijevic, L., Lindner, B., Petersen, A., Becker, W. M. and Jankov, R. M. (2008). "Quantification of the thaumatin-like kiwi allergen by a monoclonal antibody-based ELISA." Molecular Nutrition & Food Research 52(6): 701-707.
  5. Velickovic, T. C., Perovic, I., Petrovic, D., Gavrovic-Jankulovic, M., Burazer, L., Milovanovic, M., Atanaskovic-Markovic, M. and Jankov, R. (2007). "Allergenicity and immunogenicity of the major mugwort pollen allergen Art v 1 chemically modified by acetylation." Allergy 62: 260-260.
  6. N. Milosavić, R. Prodanović, R. M. Jankov; A simple and efficient one-step, regioselective, enzymatic glucosylation of arbutin by a-glucosidase Tetrahedron Letters 48(40), 7222–7224, 2007.
  7. N. Milosavić, R. Prodanović, S. Jovanović, Z. Vujčić, Immobilization of glucoamylase via its carbohydrate moiety on macroporous poly(GMA-co-EGDMA) Enzyme and Microbial Technology 40, 1422-1426, 2007.
  8. Z. Knežević, N. Milosavić, D. Bezbradica, Ž, Jakovljević, R. Prodanović; Covalent immobilization of lipase from Candida rugosa on Eupergit supports Biochemical Engineering Journal 30, 269-278, 2006.
  9. R. Prodanović, N. Milosavić, S. Jovanović, O. Prodanović, T. Čirković Veličković, Z. Vujčić, R. M. Jankov; Activity and stability of soluble and immobilized α-glucosidase from baker s yeast in cosolvent systems Biocatalysis and Biotransformation 24 (3)195-200, 2006.
This web page is supported by Faculty of Chemistry, University of Belgrade