Dynamic kinetic asymmetric enzymes

A classical approach to driving the unfavorable equilibrium of an enzymatic process is to couple it to another, irreversible enzymatic process. Griengl and coworkers have applied this concept to asymmetric synthesis of 1,2-amino alcohols with a threonine aldolase [24] (Figure 6.7). While the equilibrium in threonine aldolase reactions typically does not favor the synthetic direction, and the bond formation leads to nearly equal amounts of two diastereomers, coupling the aldolase reaction with a selective tyrosine decarboxylase leads to irreversible formation of aryl amino alcohols in reasonable enantiomeric excess via a dynamic kinetic asymmetric transformation. A one-pot, two-enzyme asymmetric synthesis of amino alcohols, including noradrenaline and octopamine, from readily available starting materials was developed [25]. 

The enzyme-catalyzed kinetic asymmetric transformation (KAT) of a diastereomeric 1 1 syn anti mixture is limited to a maximum theoretical yield of 25% of one enantiomer. This important drawback has been overcome by the combination of the actions of a ruthenium complex and a lipase in a dynamic kinetic asymmetric transformation (DYKAT), the desymmetrization of racemic or diastereomeric mixtures involving interconverting diastereomeric intermediates, implying different equilibration rates of the stereoisomers. Thus, this strategy allows the preparation of optically active diols, widely employed in organic and medicinal chemistry, as they are an important source of chiral auxiliaries and ligands and they can be easily employed as precursors of much other functionality. 

Martin-Matute, B. and Backvall, J.-E. (2004). Ruthenium- and enzymedynamic kinetic asymmetric transformation of 1,4-diols Synthesis of gamma-hydroxy ketones. /. Org. Chem., 69,9191-9195. 

Leijondahl K, Boren L, Braun R, Backvall J-E. Enzyme- and ruthenium-catalyzed dynamic kinetic asymmetric transformation of 1,5-diols. Application to the synthesis of (+)-solenop-sin A. J. Org. Chem. 2009 74 1988 1993. 

Asymmetric synthesis can refer to any process which accesses homochiral products. We will focus on asymmetric synthesis from racemic or prochiral starting materials in the presence of an enantioselective catalyst (enzyme). There are four general methodologies commonly applied kinetic resolution, dynamic kinetic resolution, deracemization and... 

Pellissier, H., Recent developments in dynamic kinetic resolution. Tetrahedron, 2008, 64, 1563-1601 Turner, N.J., Enzyme catalysed deracemisation and dynamic kinetic resolution reactions. Curr. Opin. Chem. Biol., 2004, 8, 114-119 Gmber, C.C., Lavandera, I., Faber, K. and Kroutil, W., From a racemate to a single enantiomer deracemisation by stereoinversion. Adv. Synth. Catal., 2006, 348, 1789-1805 Pellissier, H., Dynamic kinetic resolution. Tetrahedron, 2003, 59, 8291-8327 Pmnies, O. and Backvall, J.-E., Combination of enzymes and metal catalysts. A powerful approach in asymmetric catalysis. Chem. Rev., 2003, 103, 3247-3261. 

The complete transformation of a racemic mixture into a single enantiomer is one of the challenging goals in asymmetric synthesis. We have developed metal-enzyme combinations for the dynamic kinetic resolution (DKR) of racemic primary amines. This procedure employs a heterogeneous palladium catalyst, Pd/A10(0H), as the racemization catalyst, Candida antarctica lipase B immobilized on acrylic resin (CAL-B) as the resolution catalyst and ethyl acetate or methoxymethylacetate as the acyl donor. Benzylic and aliphatic primary amines and one amino acid amide have been efficiently resolved with good yields (85—99 %) and high optical purities (97—99 %). The racemization catalyst was recyclable and could be reused for the DKR without activity loss at least 10 times. 

Asymmetric Transformations by Coupled Enzyme and Metal Catalysis Dynamic Kinetic Resolution... 

Hydantoinases belong to the E.C.3.5.2 group of cyclic amidases, enzymes that catalyze the hydrolysis of hydantoins 7-11,147). Because synthetic hydantoins are accessible by a variety of chemical syntheses, including Strecker reactions, enan-tioselective hydantoinase-catalyzed hydrolysis offers an attractive and general route to chiral amino acid derivatives. Moreover, because hydantoins are easily racemized chemically or enzymatically by appropriate racemases, dynamic kinetic resolution with potential 100% conversion and complete enantioselectivity is theoretically possible. Indeed, a number of such cases have been reported 147). However, if asymmetric induction is poor or if inversion of enantioselectivity is desired, directed evolution can come to the rescue. Such a case has been reported, specifically in the production of L-methionine as part of a whole cell system E. coll) (Figure 22) 148). 

The integration of a catalyzed kinetic enantiomer resolution and concurrent racemization is known as a dynamic kinetic resolution (DKR). This asymmetric transformation can provide a theoretical 100% yield without any requirement for enantiomer separation. Enzymes have been used most commonly as the resolving catalysts and precious metals as the racemizing catalysts. Most examples involve racemic secondary alcohols, but an increasing number of chiral amine enzyme DKRs are being reported. Reetz, in 1996, first reported the DKR of rac-2-methylbenzylamine using Candida antarctica lipase B and vinyl acetate with palladium on carbon as the racemization catalyst [20]. The reaction was carried out at 50°C over 8 days to give the (S)-amide in 99% ee and 64% yield. Rather surpris-... 

M. J. Kim, Y. Ahn, and J. Park, Dynamic kinetic resolutions and asymmetric transformations by enzymes couples with metal catalysis, Curr. Opin. Biotechnol. 2002, 13, 578-587. 

Kinetic resolutions in general are regularly applied in organic synthesis. Since enzymes are highly attractive for asymmetric synthesis, various types of biocatalysts have been used in enzymatic (dynamic) kinetic resolutions, but the focus will remain on lipase- and esterase-mediated resolutions as the most common tools in early steps of natural product syntheses. 

Dynamic kinetic resolution of racemic ketones proceeds through asymmetric reduction when the substrate does racemize and the product does not under the applied experimental conditions.29 For example, baker s yeast reduction of (/ /5)-2-(4-methoxyphenyl)-l,5-benzothiazepin-3,4(2H,5H)-dione gave only (25, 35)-alcohol as a product out of four possible isomers as shown in Figure 28 (a).29a Only (5)-ketone was recognized by the enzyme as a substrate and reduction of the ketone proceeded enantioselectively. The resulting product was used for the synthesis of (25, 35)-Diltiazem, a coronary vasodilator. 

A method that has been used to approach 100% theoretical yield in asymmetric syntheses is dynamic kinetic resolution, or DKR. Although this method has been practiced based on strictly chemical reactions, only those chemoenzymatic DKR reactions will be discussed here. Most often, the enzyme used by this method is a hydrolase (lipase, esterase, protease), but other enzymes such as hydantoinases, /V-acylamino acid racemases, and dehydrogenases have also been exploited to effectively carry out DKR reactions.196 For additional details the reader is directed to the many review articles written on DKR.197 206... 

Huerta FF, Minidis ABE, Backvall JE (2001) Racemisation in asymmetric synthesis. Dynamic kinetic resolution and related processes in enzyme and metal catalysis. Chem Soc Rev 30 321-331... 

The most powerful approaches, which can be used with several different enzyme systems, lead to a single enantiomer as the product in high yield and do not rely on a classic resolution approach in which the unwanted enantiomer is discarded. These approaches include dynamic kinetic resolutions, der-acemizations, and asymmetric and desymmetrization reactions (49, 50). In some cases, a chemical catalyst may be available to recycle the unwanted isomer in the same reactor vide infra). It is sometimes possible to racemize the unwanted isomer of the substrate and then to perform the reaction again (51). 

Martin-Matute B, Backvall JE (2008) Dynamic kinetic resolutions. In Gotor V, Affonso I, Garcia-Urdiales E (eds) Asymmetric organic synthesis with enzymes. Wiley-VCH, Weinheim, pp 89-113... 

A dynamic kinetic resolution extends the high yield advantage of desymmeUiza-tions to racemic substrates. A dynamic kinetic resolution is a kinetic resolution combined with rapid in situ racemization of the substrate. The requirements for a dynamic kinetic resolution are (1) the substrate must racemize at least as fast as the subsequent enzymatic reaction, (2) the product must not racemize, and (3) as in any asymmetric synthesis, the enzymic reaction must be highly stereoselective. The equations relating product enantiomeric purity and enantioselectivity are the same as those for desymmetrizations. 

Kim, M.-J., Ahn, Y., Park, J. Dynamic kinetic resolution and asymmetric transformations by enzyme-metal combination. In Biocatalysis in the Pharmaceutical and Biotechnology Industries (ed. Patel, R.N.), 2007, CRC Press, Boca Raton, FL, 249-272. 

Transaminases can either be uhlized in kinetic resolution or as)unmetric synthesis (Scheme 29.3). Asymmetric synthesis, starting with a prochiral ketone substrate, can theoretically lead to 100% conversion and is usually the preferred route to chiral products (Scheme 29.3a). Furthermore high enantiomeric purity is not dependent on conversion rates, whereas a kinetic resolution (Scheme 29.3b) needs 50% conversion for a high enantiomeric excess (ee). But kinetic resolution is thermodynamically favored, if pyruvate is the amino acceptor, compared to as5munetric synthesis where the equilibrium lies on the substrate side [5,34]. To achieve 100% conversion, dynamic kinetic resolution serves as an alternahve with spontaneous deracemization or the initiation of a suitable racemate for enantiomerically pure substrates (Scheme 29.3c). Deracemization in a one-pot two-step reaction with an (S)-and (R)-selective transaminase, respectively, is a method of choice, but unfortunately two enantiocomplemen-tary enzymes are needed (Scheme 29.3d) [35]. Therefore deracemization with a dehydrogenase in the kinetic resolution step and a transaminase in the following step... 

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