Peptide synthesis catalyzed by an antibody containing a binding site for variable amino acids

Science

Volumn 265, Issue 5169, 1994, Pages 234-237

  Hirschmann, Ralph ^a   Smith III, Amos B ^a   Taylor, Carol M ^a   Benkovic, Patricia A ^b   Taylor, Scott D ^b   Yager, Kraig M ^a   Sprengeler, Paul A ^a   Benkovic, Stephen J ^b  

^a UNIVERSITY OF PENNSYLVANIA   (United States)

^b PENNSYLVANIA STATE UNIVERSITY   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

HAPTEN; LEUCINE; MONOCLONAL ANTIBODY; PHENYLALANINE; PHOSPHONIC ACID ESTER; TRYPTOPHAN DERIVATIVE; VALINE;

AMINO ACID ANALYSIS; ANTIBODY COMBINING SITE; ARTICLE; CATALYSIS; PEPTIDE SYNTHESIS; PRIORITY JOURNAL; PROTEIN PROTEIN INTERACTION; RACEMIC MIXTURE;

EID: 0028131354     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.8023141     Document Type: Article

References (32)

1. R. Schwyzer and P. Sieber, Helv. Chim. Acta 49, 134 (1966).
2. R. B. Merrifield, J. Am. Chem. Soc. 85, 2149 (1963); B. Gutte and R. B. Merrifield, ibid 91, 501 (1969).
3. R. B. Merrifield, J. Am. Chem. Soc. 85, 2149 (1963); B. Gutte and R. B. Merrifield, ibid 91, 501 (1969).
4. G. Siepke, H. Mullner, U. Grau, Angew. Chem. Int. Ed. Engl. 25, 535 (1986).
5. E. T. Kaiser et al., Science 243, 187 (1989); J. Rivier, J. Chromatogr. 1, 343 (1978); C. Hoeger, J. Porter, J. Boublik, J. Rivier, ibid. 404, 307 (1989); D. A. Kirby, C. L. Miller, J. E. Rivier, ibid. 648, 257 (1993).
6. E. T. Kaiser et al., Science 243, 187 (1989); J. Rivier, J. Chromatogr. 1, 343 (1978); C. Hoeger, J. Porter, J. Boublik, J. Rivier, ibid. 404, 307 (1989); D. A. Kirby, C. L. Miller, J. E. Rivier, ibid. 648, 257 (1993).
7. E. T. Kaiser et al., Science 243, 187 (1989); J. Rivier, J. Chromatogr. 1, 343 (1978); C. Hoeger, J. Porter, J. Boublik, J. Rivier, ibid. 404, 307 (1989); D. A. Kirby, C. L. Miller, J. E. Rivier, ibid. 648, 257 (1993).
8. E. T. Kaiser et al., Science 243, 187 (1989); J. Rivier, J. Chromatogr. 1, 343 (1978); C. Hoeger, J. Porter, J. Boublik, J. Rivier, ibid. 404, 307 (1989); D. A. Kirby, C. L. Miller, J. E. Rivier, ibid. 648, 257 (1993).
9. R. Hirschmann et al., J. Am. Chem. Soc. 91, 509 (1969).
10. H.-D. Jakubke, in The Peptides, S. Udenfriend and J. Meienhofer, Eds. (Academic Press, San Diego, CA, 1987), vol. 9, chap. 3, pp. 103-159. An attractive molecular biology-based approach has recently been described [L. Arahmsér et al., Biochemistry 30, 4151 (1991)].
11. H.-D. Jakubke, in The Peptides, S. Udenfriend and J. Meienhofer, Eds. (Academic Press, San Diego, CA, 1987), vol. 9, chap. 3, pp. 103-159. An attractive molecular biology-based approach has recently been described [L. Arahmsér et al., Biochemistry 30, 4151 (1991)].
12. R. A. Lerner, S. J. Benkovic, P. G. Schultz, Science 252, 659 (1991); S. J. Benkovic, Annu. Rev. Biochem. 61, 29 (1992).
13. R. A. Lerner, S. J. Benkovic, P. G. Schultz, Science 252, 659 (1991); S. J. Benkovic, Annu. Rev. Biochem. 61, 29 (1992).
14. Although ours is the first documented, P. G. Schultz [Angew. Chem. Int. Ed. Engl. 28, 1283 (1989)] has referred to unpublished results for peptide bond formation.
15. Although our initial target hapten was 1, incorporating a phosphonamide unit as the mimic of the tetrahedral intermediate, phosphonate 2 proved to be more synthetically accessible. An immunogenic conjugate was then prepared from 2a and keyhole lympet hemocyanin. Monoclonal antibodies were prepared by standard protocols [G. Kohler and C. Milstein, Nature 256, 495 (1975)].
16. The 24 monoclonal antibodies were selected on the basis of their ability to bind a bovine serum albumin conjugate with 2 and were purified to >90% homogeneity, as judged by SDS-gel electrophoresis. All 24 antibodies were screened for their ability to catalyze the coupling of the p-chlorophenyl, p-nitrobenzyl (2.0 mM), or p-nitrophenyl (1.0 mM) esters of N-acetyl-L-valine (3, 4, and 5a, respectively) with 2.0 mM D-tryptophan amide 6 at pH 8.3 (3 and 4) or pH 7.0 (5a) to form dipeptide 7a.
17. 18 reversed-phase column (VYDAC, Hesperia, CA) and eluted with an acetonitrile-water (0.1% trifluoroacetic acid) gradient with the detector set at 280 nM. We identified the product by comparing the retention time of the product formed during the reaction with that of authentic samples. The relative rates were determined by dividing the amount of 7 formed by the antibody with that formed in the control reaction.
18. The control experiments were carried out under the same conditions as described (11), except antibody was not added to the reaction mixture.
19. A. R. Fersht and W. P. Jencks, J. Am. Chem. Soc. 92, 5442 (1970).
20. -1.
21. R. Hirschmann et al., unpublished results.
22. A. D. Napper, S. J. Benkovic, A. Tramontano, R. A. Lerner, Science 237, 1041 (1987).
23. A. G. Cochran, T. Pham, R. Sugasawara, P. G. Schultz, J. Am. Chem. Soc. 113, 6670 (1991).
24. J. R. Jacobsen, J. R. Prudent, L. Kochersperger, S. Yonkovich, P. G. Schultz, Science 256, 365 (1992). In this case, the dissociation constant of 18 nM for the product relative to a value of 0.77 mM for the alcohol substrate effectively inhibited the reaction after one turnover.
25. A. L. Heard and G. T. Young, J. Chem. Soc. 1963, 5807 (1963); N. L. Benoiton, in The Peptides, S. Udenfriend and J. Meienhofer, Eds. (Academic Press San Diego, 1983), vol. 5, chap. 4.
26. A. L. Heard and G. T. Young, J. Chem. Soc. 1963, 5807 (1963); N. L. Benoiton, in The Peptides, S. Udenfriend and J. Meienhofer, Eds. (Academic Press San Diego, 1983), vol. 5, chap. 4.
27. R. Stanfield, M. Takimoto-Kamimura, J. Rim, A. T. Profy, I. Wilson, Structure 1, 83 (1993); J. Moult et al., J. Mol. Biol. 74, 137 (1976).
28. R. Stanfield, M. Takimoto-Kamimura, J. Rim, A. T. Profy, I. Wilson, Structure 1, 83 (1993); J. Moult et al., J. Mol. Biol. 74, 137 (1976).
29. J. H. Arevalo, M. J. Tausig, I. A. Wilson, Nature 365, 859 (1993).
30. B. S. Green, in Catalytic Antibodies (CIBA Foundation Symposium 159, Wiley, West Sussex, United Kingdom, 1991), p. 139.
31. J. D. Stewart and S. J. Benkovic, Chem. Soc. Rev. 22, 213 (1993).
32. R.H. acknowledges J. R. Huff, E. Thornton, and T. Widlanski for substantive discussions. We thank D. Schloeder for assistance in the preparation of the monoclonal antibodies and B. Jiang for assisting S.D.T. and P.A.B. Supported by NIH (Institute of General Medical Sciences) through grant GM-45611 (R.H. and A.B.S.). Bachem (R.H.), and Merck Research Laboratories (R.H.). C.M.T. is a recipient of a William Georgetti Scholarship. S.D.T. is a recipient of a Natural Sciences and Engineering Research Council of Canada postdoctoral fellowship. K.M.Y. is a recipient of an NIH (National Cancer Institute) postdoctoral fellowship.