Novel enantioselective enzymes from non-conventional yeasts

The major goal of my research is the development of synthetically useful selective biotransformations using “new” microbial enzymes by combining natural and non-natural conditions. In this poster will be presented results from research activities on three different class of enzymes: ● Carboxylesterases – A new cell-bound carboxylesterase from Kluyveromyces marxianus have been purified and partially characterised. It has been employed for different enantioselective hydrolysis of chiral esters (i.e. solketal and aminoacid esters), not easily and/or efficiently resolved with commercial enzymes. Different strategies of optimisation and development of processes yelding multi-gram scale production will be also discussed. Ref. Daniela Monti, Erica Elisa Ferrandi, Monica Righi, Diego Romano, Francesco Molinari. Purification and characterization of the enantioselective esterase from Kluyveromyces marxianus CBS 1553. Journal of Biotechnology (2008), 133(1), 65-72. ● Dehydrogenases - Pichia glucozyma produces DHs able to reduce aromatic ketones with peculiar stereodifferentiation between the prostereogenic faces of the carbonyls. Different aromatic substrates were reduced with high molar conversions and excellent stereoselectivity; the stereoselectivity was often complementary to the one observed with organometallics. The presence of different dehydrogenases in Pichia glucozyma allowed also for the deracemization of different racemic benzoins and α-substituted carbonyls. Ref. Syed Masood Husain, Thomas Stillger, Pascal Dünkelmann, Melanie Lödige, Lydia Walter, Elke Breitling, Martina Pohl, Mara Bürchner, Ingo Krossing, Michael Müller, Diego Romano, Francesco Molinari, Stereoselective reduction of 2-hydroxy ketones towards syn- and anti-1,2-Diols – Accepted for publication in Advanched Sinthesys and Catalisys R. Gandolfi, E. Cesarotti, F. Molinari, D. Romano, I. Rimoldi. Asymmetric reductions of ethyl 2-(benzamidomethyl)-3-oxobutanoate by yeasts Tetrahedron: Asymmetry 2009, 20, 411-414. ● Enoate Reductases (OYEs) - Old Yellow Enzymes are flavin-dependent oxidoreductases that catalyze the stereoselective trans-hydrogenation of electron-poor alkenes (enoate reductase activity), which can result in important chiral synthons toward synthesis of many pharmaceuticals and chemicals. In this study, environmental yeasts have been screened for OYE activity with the purpose to develop new enoate reductase biocatalyst. Recombinant strains with enhanced OYE activity have been obtained and could be exploited for production of purified enzyme. Furthermore, the data herein reported attract interest due to promising applications in asymmetric synthesis. Ref. Stefano Raimondi, Diego Romano, Alberto Amaretti, Francesco Molinari and Maddalena Rossi. Enoate reductases from non conventional yeasts as novel biocatalysts: cloning and functional expression in Saccharomyces cerevisiae. Submitted at Journal of Biotechnology