VOLUME117, NUMBER 27 JULY 12, 1995 0Copyright 1995 by the American Chemical Society
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY Anti-Metallocene Antibodies: A New Approach to Enantioselective Catalysis of the Diels- Alder Reaction4 Jari T. Yli-Kauhaluoma,+Jon A. Ashley, Chih-Hung Lo, Lee Tucker, Mary M. Wolfe, and Kim D. Janda* Contribution from the Departments of Molecular Biology and Chemistry, The Scripps Research Institute, 10666 North Torrey Pines Road, La Jolla, Califomia 92037 Received February I , 1995@
Abstract: We have shown how a constrained bicyclo[2.2.2]octene hapten can elicit antibody catalysts for the DielsAlder reaction between 4-carboxybenzyl trans- 1,3-butadiene-1-carbamate and NJV-dimethylacrylamide(Gouvemeur, V. E.; de Pascual-Teresa, B.; Beno, B.; Janda, K. D.; Lemer, R. A. Science 1993, 262, 204). In the present study we have developed a new approach to hapten design for elicitation of Diels-Alder catalytic antibodies. Our strategy was to engage the freely rotating $-cyclopentadienyl iron complex as the haptenic group. By applying such a flexible hapten we set out to determine if the immune system could freeze out a conformer which mimics the DielsAlder transition state and hence produce new Diels-Alderases. If so, how would the catalytic rates, diastereo- and enantioselectivity of these antibodies compare with those of antibodies elicited by the former methodology? We generated antibodies that catalyzed the Diels-Alder reaction with high enantio- and diastereoselectivity and had effective molarities (EM) comparable to those of antibodies elicited using the constrained bicyclo[2.2.2]octene haptens. This methodology offers a new approach to the production of antibodies for the catalysis of other reactions with pericyclic, highly-ordered transition states.
Introduction In view of its synthetic utility and the lack of a known biological catalyst, the Diels- Alder reaction has been seen as an important focus for the catalytic antibody field.’-3 A number of groups, including our own, have successfully generated antibodies capable of catalyzing Diels-Alder cycloaddition rea~tions.4,~ Our initial approach, using the bicyclo[2.2.2]octene haptens 1 and 2 (Figure l), was to induce antibody catalysts which could individually control the product distribution of the reaction between diene 3 and dienophile 4.6 The selection of
* Author to whom correspondence should be addressed.
’ On leave from The Technical Research Centre of Finland, Chemical
Technology, Biologinkuja 7, FIN-02 150 Espoo, Finland. Abbreviations: EDJA, Nb”diisopropylethy1amine; TEA, triethylamine: t-BOC. rert-butoxycarbonyl; BOPCl. bis(2-oxo-3-oxazolidinyl)phosphinic chloride: 3-NBA, 3-nitrobenzyl alcohol: PMA. phosphomolybdic acid: DSS, sodium 2.2-dimethyl-2-silapentan-5-sulfonate; EDC, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimidehydrochloride. Abstract published in Advance ACS Abstracts. June 15, 1995. 7
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these highly constrained structures (1 and 2) was based on elegant work previously reported by Schultz$b who hypothesized that the bicyclo[2.2.2]octene system might be a good mimic for the ordered transition state of the Diels-Alder reaction. This was later confirmed by ab initio calculation^.^ (1) (a) Diels, 0.:Alder, K. Liebigs Ann. Chem. 1928,460.98- 122. (b) Canuthers, W. Cycloaddition Reactions in Organic Chemist?: Tetrahedron Organic Chemistry Series Volume 8: Pergamon Press: Oxford, 1990. (c) Oppolzer, W. Intermolecular Diels- Alder reactions. In Comprehensive Organic Synthesis:Trost, B. M., Fleming, I., Paquette, L. A,. Eds.: Pergamon Press: Oxford. 1991: Vol. 5, pp 315-399. (d) Oppolzer. W. Angew. Chem., lnt. Ed. Engi. 1977, 16. 10-23. (e) Pindur, U.: Lutz. G.: Otto. C. Chem. Rev. 1993. 93, 741-761. (0 Oppolzer, W. Angew. Chem.. Int. Ed. Engl. 1984, 23, 876-889. (g) Kagan, H. B.: Riant, 0. Chem. Rev. 1992, 92. 1007-1019. (h) Narasaka, K. Synthesis 1991, 1-11. (2) For biocatalytic methods in the Diels-Alder reactions. see for example: (a) Colonna. S.: Manfredi. A,: Annunziata. R. Tetrahedron Lett. 1988. 29. 3347-3350. (b) Rama Rao. K.; Srinivasan, T. N.: Bhanumathi, N. Tetrahedron Lett. 1990, 31, 5959-5960. The use of bovine serum albumin and baker’s yeast to catalyze the Diels-Alder reaction is reported in these communications, respectively.
0002-786319511517-7041$09.00/0 0 1995 American Chemical Society
7042 J. Am. Chem. SOC.,Vol. 117. No. 27. 1995
Yli-Kauhaluoma et al. ofiho-endo
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Figure 2. Representations of the transition stales for the ortho-undo and orth-e.roDid-Alder rextions and the corresponding ferrocene Structure for these transition states. R = CONMe?. R' = NHCO.. CHdCoHrCO?H, R" = (CH?)CO:H or 4-carhoxyphenyl.
Results and Discussion 17 .m
Figure 1. Antibody-catalyzed Diels-Alder reactions and the SIIIICIUT~S of haptens 1. 2. 5. and 6 and inhihitor 8 used in the present study. R = COiCH+CO?H. R = 4-carhoxyhenzyl.
In our new strategy, we intended to challenge the notion that the best haptens for reactions proceeding through highly ordered transition states must be conformationally restricted analogs. Rather. we wondered how the immune system would "recognize" a conformationally unrestricted hapten. W e were interested in using the dicyclopentadienyl system of ferrocene as a loose mimic for the cyclic, six-membered transition state of the Diels- Alder reaction. Clearly, from the vast immune library, antibodies to all stable conformers of the molecule should exist. However, critical to our success would be antibody recognition and the freezing out of a conformer which resembled the DielsAlder transition state. Therefore, this conformationally flexible haptenic design offered an interesting, antithetical approach to antibody-catalysis of the Diels-Alder reaction. 131 There is romr indirect expenmental evidence for a hiolopical DielsAlder reaction imolving diene and dienophile precursors in the biosynthesis of solanapyrones although rhe isdarton and characterization of a natural Dels-Aldernse is yet to he described: in1 Oikaws. H.: Suzuki. Y.: Naga. A.: Kntaysma. ti.: Ichihnra. A. 1.A m Chrrn. .%K 1994. 116. 3605-3606. Ihl Oikawa. H.: Yokora. T.: Abe. T.: Ichihara. A.: Siikamum. S.: Yoshizawa. Y.:Vederas. I. C . J. C l m Soc.. ~ Chum. Cmmun. 1989. 1282-1284. i c ) Oikann. H.: Yokora. T.: Ichihnra. A,: Sakamurn. S. .I. Cliem. Sm.. Chm,. Commen. 1989. 1284- 1285. For several examples of postulated hiosynthetic Did-Alder adducrs. see for example: in1 Sanz-Cemera.J. F.: Glinkn. T.: Williams. R. M.J. An?. Chum SO
