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Enzyme-assisted physicochemical enantioseparation processes - part I: Production and characterization of a recombinant amino acid racemase

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http://pubman.mpdl.mpg.de/cone/persons/resource/persons86430

Petrusevska,  K.
Physical and Chemical Foundations of Process Engineering, Max Planck Institute for Dynamics of Complex Technical Systems, Max Planck Society;

http://pubman.mpdl.mpg.de/cone/persons/resource/persons86477

Seidel-Morgenstern,  A.
Physical and Chemical Foundations of Process Engineering, Max Planck Institute for Dynamics of Complex Technical Systems, Max Planck Society;
Otto-von-Guericke-Universität Magdeburg, External Organizations;

http://pubman.mpdl.mpg.de/cone/persons/resource/persons86282

Elsner,  M. P.
Physical and Chemical Foundations of Process Engineering, Max Planck Institute for Dynamics of Complex Technical Systems, Max Planck Society;

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Citation

Würges, K., Petrusevska, K., Serci, S., Wilhelm, S., Wandrey, C., Seidel-Morgenstern, A., et al. (2009). Enzyme-assisted physicochemical enantioseparation processes - part I: Production and characterization of a recombinant amino acid racemase. Journal of Molecular Catalysis B-Enzymatic, 58(1-4), 10-16. doi:10.1016/j.molcatb.2008.10.006.


Cite as: http://hdl.handle.net/11858/00-001M-0000-0013-9363-C
Abstract
The demand for enantiopure substances, e.g. for pharmaceutical applications or fine chemical production, continues to increase. This has led to the development of numerous stereoselective synthesis methods. Nevertheless a large number of chemical syntheses still result in racemic mixtures making a subsequent enantioseparation step necessary and thus are restricted to a maximum yield of 50%. Our work focuses on strategies to overcome this limitation by combining physicochemical separation processes with enzymatic racemization of the unwanted enantiomer in order to produce enantiopure amino acids. This paper deals with the production and characterization of a suitable amino acid racemase with broad substrate specificity (EC 5.1.1.10) from Pseudomonas putida which we cloned into Escherichia coli. Two enzyme lyophilizates of different purity were obtained from which the crude (CL) was sufficient for the racemization of methionine (Met) and the pure (PL) was used for asparagine (Asn). Racemization reactions of D-/L-Asn in H2O and D-/L-Met in 95 vol.% 100 mM KPi-buffer, 5 vol.% methanol (MeOH) at different pH values and temperatures were characterized. The studied range of reaction parameters was chosen in dependency on planned enantioseparation processes. We found increasing Vmax values when temperature was risen stepwise from 20 to 40 °C for both systems and when pH was shifted from 6 to 8 for the Met system. The presented results provide the basis for engineering enzyme-assisted physicochemical enantioseparation processes. Copyright © 2008 Elsevier B.V. All rights reserved. [accessed May 11, 2009]