An aspartate and a water molecule mediate efficient acid-base catalysis in a tailored antibody pocket

Erik W. Debler; Roger Müller; Donald Hilvert; Ian A. Wilson

doi:10.1073/pnas.0902700106

New Research In

Edited by Richard Wolfenden, University of North Carolina, Chapel Hill, NC, and approved September 11, 2009 (received for review March 11, 2009)

Abstract

Design of catalysts featuring multiple functional groups is a desirable, yet formidable goal. Antibody 13G5, which accelerates the cleavage of unactivated benzisoxazoles, is one of few artificial enzymes that harness an acid and a base to achieve efficient proton transfer. X-ray structures of the Fab-hapten complexes of wild-type 13G5 and active-site variants now afford detailed insights into its mechanism. The parent antibody preorganizes Asp^H35 and Glu^L34 to abstract a proton from substrate and to orient a water molecule for leaving group stabilization, respectively. Remodeling the environment of the hydrogen bond donor with a compensatory network of ordered waters, as seen in the Glu^L34 to alanine mutant, leads to an impressive 10⁹-fold rate acceleration over the nonenzymatic reaction with acetate, illustrating the utility of buried water molecules in bifunctional catalysis. Generalization of these design principles may aid in creation of catalysts for other important chemical transformations.

Footnotes

³To whom correspondence may be addressed. hilvert{at}org.chem.ethz.ch or wilson{at}scripps.edu
Author contributions: E.W.D., R.M., D.H., and I.A.W. designed research; E.W.D. and R.M. performed research; E.W.D. and R.M. contributed new reagents/analytic tools; E.W.D., R.M., D.H., and I.A.W. analyzed data; and E.W.D., R.M., D.H., and I.A.W. wrote the paper.
The authors declare no conflict of interest.
This article is a PNAS Direct Submission.
Data deposition: The atomic coordinates and structure factors have been deposited in the Protein Data Bank, www.pdb.org [PDB ID codes 3FO0 (hapten complex of the 13G5 wild-type), 3FO1 (hapten complex of the 113G5-Glu^L34Ala variant), and 3FO2 (hapten complex of the 13G5-Glu^L34Gln variant)].

Erik W. Debler^a,¹,
Roger Müller^b,²,
Donald Hilvert^b,³ and
Ian A. Wilson^a,³

^aDepartment of Molecular Biology and Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037 and
^bLaboratorium für Organische Chemie, ETH Zürich, CH-8093 Zürich, Switzerland

↵¹Present address: Laboratory of Cell Biology, Howard Hughes Medical Institute, 1230 York Avenue, Rockefeller University, New York, NY 10065.
↵²Present address: Munich Center for Integrated Protein Science (CIPSM) at the Department Chemie, Technische Universität München, D-85747 Garching, Germany.

Edited by Richard Wolfenden, University of North Carolina, Chapel Hill, NC, and approved September 11, 2009 (received for review March 11, 2009)

Abstract

catalytic antibody
crystal structure
enzyme design
enzyme mechanism
proton transfer

Footnotes

³To whom correspondence may be addressed. hilvert{at}org.chem.ethz.ch or wilson{at}scripps.edu
Author contributions: E.W.D., R.M., D.H., and I.A.W. designed research; E.W.D. and R.M. performed research; E.W.D. and R.M. contributed new reagents/analytic tools; E.W.D., R.M., D.H., and I.A.W. analyzed data; and E.W.D., R.M., D.H., and I.A.W. wrote the paper.
The authors declare no conflict of interest.
This article is a PNAS Direct Submission.
Data deposition: The atomic coordinates and structure factors have been deposited in the Protein Data Bank, www.pdb.org [PDB ID codes 3FO0 (hapten complex of the 13G5 wild-type), 3FO1 (hapten complex of the 113G5-Glu^L34Ala variant), and 3FO2 (hapten complex of the 13G5-Glu^L34Gln variant)].