Antibody catalyzed Diels-Alder reaction

Catalytic antibody 1E9, the first catalytic antibody discovered for Diels-Alder reaction, catalyzes the cycloaddition between tetrachlorothiophene dioxide and N-ethylmaleimide (Equation 4.15) [86]. 

Yli-Kauhaluoma J. Control of Regioselectivity, Diastereoselectivity and Enantios-electivity in the Antibody-Catalyzed Diels-Alder Reaction VTT Symp. 1996 163 69-74... 

Keywords antibodies that catalyze the Diels-Alder reaction... 

Keywords stable analogs of transition state are used as haptens to elicit antibodies that will catalyze Diels-Alder reaction... 

In addition to Lewis acids, Diels-Alder reactions in aqueous media are also catalyzed by bovine serum albumin,49 enzymes,50 antibodies,51 and amines.52... 

The antibody-catalyzed Diels-Alder reaction developed by Schultz utilized a Diel-Alderase enzyme-like catalyst evolved from an antibody-combining site (Eq. 12.13). The idea is that the generation of antibodies to a structure that mimics the transition state for the Diels-Alder reaction should result in an antibody-combining site that lowers the entropy of activation by binding both the diene and dienophile in a reactive conformation. 

Similar aza-Diels-Alder reactions of Danishefsky s diene with imines or aldehydes and amines in water took place smoothly under neutral conditions in the presence of a catalytic amount of an alkaline salt such as sodium triflate or sodium tetraphenylborate to afford dihydro-4-pyridones in high yields (Eq. 12.49).117 Antibodies have also been found to catalyze hetero-Diels-Alder reactions.118... 

The Diels-Alder reaction is one of the most powerful and versatile carbon-carbon bond-forming methods available to synthetic organic chemists attempts to isolate enzymes that catalyze such a process have been unsuccessful. Therefore, the acceleration of this reaction by an abzyme has been an important landmark in the field of catalytic antibodies and of considerable potential for chemical synthesis. 

Figure 8 Diels-Alder reaction catalyzed by the antibody 1E9 raised against endo-hexachloronorbornene 7 and X-ray...

Figure 9 Diels-Alder reaction catalyzed by antibody 39A11 raised against the bicyclic hapten 8.

While indirect selections work quite well for antibodies they have been less successful in the case of catalytic nucleic acids. There are only three examples which prove that it is possible in principle to obtain a ribo- or deoxyribozyme by selecting an aptamer that binds to a TSA A rotamase ribozyme [7], a ribozyme capable of catalyzing the metallation of a porphyrin derivative [92], and one catalytic DNA of the same function [93]. Another study reported the selection of a population of RNA-aptamers which bind to a TSA for a Diels-Alder reaction but the subsequent screen for catalytic activity was negative for all individual RNAs tested [94]. The attempt to isolate a transesterase ribozyme using the indirect approach also failed [95]. 

Recently, an antibody has been described which catalyzes not just a Diels-Alder reaction between an N-substituted maleimide and acetoxybutadiene (kcat = 0.055 s-1, Km = S3 mM, kcat/fCM = 6.6 (m s) but also the subsequent hydrolysis oftheacetoxy group (kcat = 9.2 x 1(T4 s 1, JCM = 1.1 rmvr, kcat/KM = 0.9 (m s) 1), which is about 1.5% as fast as the Diels-Alder reaction itself (Figure 18.9). 

Figure 18.7 Diels-Alder reactions catalyzed by antibodies (Lerner, 1991).

Pandit and co-workers adopted the same strategy to catalyze a hetero-Diels-Alder reaction in which an aryl-nitroso derivative serves as the dienophile.117 118 Antibodies generated against thebicyclic transition-state analogs 133 and 134 accelerated the reaction between the trans-diene 135 and 136, but the ratio of the two product regioisomers 137 and 138 was the same as for the uncatalyzed reaction (58 42). In experiments with stoichiometric amounts of trans-135 and antibody 309-1G7, derived from hapten 134, this ratio was altered somewhat in favor of product 138 (47 53), in accord with the structure of the... 

Scheme 4.3 Antibody 39-All catalyzes a Diels—Alder reaction between an electron-rich acyclic diene (8) and an N-aryl maleimide (9). It was elicited with the bicyclo[2.2.2]octene hapten 7. The ethano bridge locks the cyclohexene ringintothe requisite boat conformation but has no counterpart in the substrates or transition state.

The question whether Diels-Alder reactions also occur in nature can not be answered yet since special enzymes catalyzing these reactions have not been found so far [24-25]. However, artificial catalytic antibodies for Diels-Alder reactions are well known [26] and recently an all-carbon [4 + 2]cycloaddition has been observed in the biosynthesis of two phytotoxic solanopyrones 1-1 and 1-2 from the fungus Alternaria solani using a cell-free extract of this organism [27] it is highly probable that the involved enzymes will soon be isolated (Fig. 1-2). 

Scheme 3. The highly-ordered transition state associated with the Diels-Alder reaction has made it a natural target for work in the field of catalytic antibodies [15]. Indeed, Hilvert [15a] has reported recently the first successfully antibody-catalyzed Diels-Alder reaction. Monoclonal antibodies elicited to the boxed hapten 14 provided both acceleration of and multiple turnover of the cycloaddition between tetrachlorothiopene dioxide and IV-ethyl maleimide. The initial adduct decomposes by the chelotropic-extrusion of sulfur dioxide affording the dihydrophthalimide derivative 13...

Figure 10 (a) Hapten 24 elicited antibody 1E9 that catalyzes the Diels-Alder reaction between diene 25 and dienophile 26. (b) Key contacts of the 1E9 Fab-24 complex. 

Cannizzaro CE, Ashley JA, Janda KD, Houk KN. Experimental determination of the absolute enantioselectivity of an antibody-catalyzed Diels-Alder reaction and theoretical explo- 110. rations of the origins of stereoselectivity. J. Am. Chem. Soc. 2003 125(9) 2489-2506. 

Cannizzaro, C. E., Ashley, J. A., Janda, K. D., Houk, K. N. Experimental Determination of the Absolute Enantioselectivity of an Antibody-Catalyzed Diels-Alder Reaction and Theoretical Explorations of the Origins of Stereoselectivity. J. Am. Chem. Soc. 2003,125, 2489-2506. 

Antibodies can be utifized under a variety of non-physiological conditions and are excellently suited for preparative applications due to their intrinsic stability. We have found that antibodies catalyzing the retro-Diels-Alder reaction of 63 function equally well between pH 4 and pH 11. Aldolase antibody 72D4 operates in the presence of 10% acetone. Janda et al. have used an immobilized esterase antibody with up to 40% dimethylsulfoxide [110]. Esterase catalytic antibodies have been used in reverse micelles and in lipid-coated form to transform lipophilic substrates [111]. Catalytic antibodies can also be used in a biphasic alkane/water system [112]. The lipophilic substrate remains in the alkane phase where it does not undergo any reaction, which suppresses any uncatalyzed reaction. In case that the reaction product is still lipophilic and returns to the alkane phase, product inhibition is also suppressed under these conditions. 

However, complete stereochemical control of the Diels-Alder reaction to yield only disfavored exo-products in enantiomerically pure form has proven to be very difficult by chemical means. Furthermore, only recently has a potentially enzymatic Diels-Alder reaction been reported [37]. Therefore, attempts to generate antibodies which can catalyze stereoselective Diels-Alder reactions is seen as an ongoing major target in the field. 

Each antibody catalyzes its respective processes not only with high diastere-oselectivity (>98% de), but also with excellent enantioselectivity (>98% ee), such that the antibody-catalyzed Diels-Alder reaction gives essentially optically pure endo-27 or exo-28 adducts. 

There are several examples of Diels-Alder reactions catalyzed by catalytic antibodies and RNA. Their mechanisms will be discussed in this section. 

The Diels-Alder reaction usually does not require a catalyst, and therefore, rate acceleration may be achieved by stabilizing the transition state. In fact, antibodies elicited by transition state analog are able to catalyze Diels-Alder reactions. The Diels-Alderase antibodies provide a pocket acting as an entropy trap. Using this strategy several Diels-Alderase antibodies have been created to date. " Product inhibition is an inherent problem in the use of Diels-Alderase antibodies because of the resemblance between the product and the transition state. 

To escape product inhibition, unstable boat-like haptens are used to prepare Diels-Alderase antibodies. Braisted and Schultz raised antibody 39-All with hapten 124 fixed in a boat form. The antibody 39-All catalyzed the Diels-Alder reaction between 121 and 122 to yield the chiral adduct 123 (Scheme 21). 

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