Steven Boxer
Camille Dreyfus Professor
Department of Chemistry
Stanford University

Steven Boxer is the Camille Dreyfus Professor in the Department of Chemistry at Stanford University. His research interests are in biophysics: the interface of physical chemistry, biology and engineering. Topics of current interest include: electrostatics and dynamics in proteins, especially related to enzyme catalysis; excited state dynamics of green fluorescent protein, especially split GFP, with applications in biotechnology; electron and energy transfer mechanisms in photosynthesis; and the fabrication of artificial systems to simulate, manipulate and image biological membranes. He has served on the scientific advisory board of many start-ups in the general area of biotechnology, and as an advisor to government and non-profit organizations in the U.S. and around the world. He is the recipient of several awards, and is an elected Fellow of the American Academy of Arts and Sciences, the Biophysical Society and the National Academy of Sciences.

Electric Fields and Enzyme Catalysis

The origin of the remarkable rate acceleration exhibited by enzymes is a topic of longstanding debate and many ideas have been proposed. Electrostatic interactions impact every aspect of the structure and function of proteins, nucleic acids, and membranes. The transition states for many enzyme-catalyzed reactions involve a change in the distribution of charge relative to the starting material and/or products, and the selective stabilization of charge-separated transition states may be essential for catalysis. The magnitudes of the electric fields in proteins and the variations in these fields at different sites are predicted to be enormous, but it is a challenge to obtain quantitative experimental information on these fields. We have developed the vibrational Stark effect to probe electrostatics and dynamics in organized systems, in particular in proteins where they can report on functionally important electric fields. The strategy involves deploying site-specific vibrational probes whose sensitivity to an electric field is measured in a calibrated external electric field. Once calibrated, these probes, typically nitriles or carbonyls, can be used to probe changes in electric field due to mutations, ligand binding, pH effects, light-induced structural changes, etc. We can also obtain information on absolute fields by combining vibrational solvatochromism and MD simulations, checked by the vibrational Stark effect calibration. This frequency-field calibration can be applied to quantify functionally relevant electric fields at the active site of enzymes. Using ketosteroid isomerase (KSI) as a model system, we correlate the field sensed at the bond involved in enzymatic catalysis with the rate of the reaction it catalyzes, including variations in this rate in a series of mutants and variants using non-canonical amino acids. This provides the first direct connection between electric fields and function: for this system electrostatic interactions are a dominant contribution to catalytic proficiency. Using the vibrational Stark effect, we can now consistently reinterpret results already in the literature and provide a framework for parsing the electrostatic contribution to catalysis in both biological and non-biological systems. Electric fields provide a physics-based metric for the origin of function and progress towards connecting this metric with the evolutionary history of enzymes will be discussed.