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Steven Boxer
Camille Dreyfus Professor of Chemistry
Department:
Chemistry
PhD, University of Chicago, Physical and Physical-Organic Chemistry (1976)
BS with Honors, Tufts University, Chemistry (1969)
My laboratory investigates the structure and function of biological systems with a strong physical perspective. We invent experimental methods and develop theory as needed. We are pursuing several interconnected themes:
Electrostatics and Dynamics in Proteins
We study electrostatics in proteins and how electric fields affect function. Early test systems used mutants of myoglobin, which was first cloned and expressed in our lab. This led to probes whose sensitivity to electric fields can be calibrated by Stark spectroscopy — spectroscopy in electric fields — which we have developed into a broadly applicable method. Vibrational Stark experiments exploit molecular vibrations as local and directional probes to map electrostatic fields in proteins. These probes include nitriles (CN), carbonyls (C=O) and C-D. These can be used to probe electrostatics and to make direct comparisons and tests of simulations. Recent work focuses on carbonyl probes to study enzymatic reactions. By combining the vibrational Stark effect, vibrational solvatochromism and MD simulations, we have developed a general method to measure the absolute field sensed by the carbonyl probe in proteins. This has been used to quantify the electrostatic contribution to the catalytic rate in several enzymes.
Excited State Dynamics in GFP and Quantum Biology
Green Fluorescent Protein (GFP) is widely used as a probe to localize proteins in cells. Our lab was the first to demonstrate that the GFP chromophore exists in two protonation states, interconvertible by ultrafast excited state proton transfer. We and others have since developed this idea to generate novel GFP variants with diverse colors and sensitivities. The GFP chromophore has proven to be a sensitive probe for electrostatics inside the proteins. Modifications of the chromophore using non-canonical amino acids and environmental perturbations have been used to probe the role of electrostatics in chromophore photoisomerization. The discovery that fluorescence from red fluorescent proteins (RFPs) in the presence of flavins depends on external magnetic fields has returned our lab to earlier work on magnetic field effects that arise from spin-correlated radical pairs. Understanding the origin(s) of these effects may help to develop these effects as an imaging method.
Model Membranes
Our group has developed supported lipid bilayers as mimics for cell surfaces and tools in biotechnology. A broad vision is to engineer interfaces between hard surfaces and soft materials, ultimately leading to sophisticated biocompatible interfaces that can be used to control, interrogate or organize complex living systems. We have developed methods to partition and manipulate elements of these unique self-assembled systems; these methods are now used in many laboratories.
Recent work addresses three interrelated areas: 1) characterization of membrane organization, domains and protein associations using a novel type of imaging mass spectrometry; 2) models for membrane fusion and investigations into the fusion of enveloped viruses and lipid nanoparticles to their target membrane; 3) development of tethered lipid bilayers as a platform to study membrane domains and junction topology and organization between supported membranes and natural cell membranes.
Energy and Electron Transfer in Photosynthesis
Light-driven long-distance electron transfer in photosynthetic reaction centers is one of the fastest known chemical reactions. We studied this by femtosecond fluorescence and transient absorption spectroscopy, manipulation in electric fields, site-specific and global mutagenesis and some novel types of Stark spectroscopy. While this phase of our work is complete, we continue to use elements of the photosynthetic apparatus to study the electric fields in membranes created in response to transmembrane potential.
Electrostatics and Dynamics in Proteins
We study electrostatics in proteins and how electric fields affect function. Early test systems used mutants of myoglobin, which was first cloned and expressed in our lab. This led to probes whose sensitivity to electric fields can be calibrated by Stark spectroscopy — spectroscopy in electric fields — which we have developed into a broadly applicable method. Vibrational Stark experiments exploit molecular vibrations as local and directional probes to map electrostatic fields in proteins. These probes include nitriles (CN), carbonyls (C=O) and C-D. These can be used to probe electrostatics and to make direct comparisons and tests of simulations. Recent work focuses on carbonyl probes to study enzymatic reactions. By combining the vibrational Stark effect, vibrational solvatochromism and MD simulations, we have developed a general method to measure the absolute field sensed by the carbonyl probe in proteins. This has been used to quantify the electrostatic contribution to the catalytic rate in several enzymes.
Excited State Dynamics in GFP and Quantum Biology
Green Fluorescent Protein (GFP) is widely used as a probe to localize proteins in cells. Our lab was the first to demonstrate that the GFP chromophore exists in two protonation states, interconvertible by ultrafast excited state proton transfer. We and others have since developed this idea to generate novel GFP variants with diverse colors and sensitivities. The GFP chromophore has proven to be a sensitive probe for electrostatics inside the proteins. Modifications of the chromophore using non-canonical amino acids and environmental perturbations have been used to probe the role of electrostatics in chromophore photoisomerization. The discovery that fluorescence from red fluorescent proteins (RFPs) in the presence of flavins depends on external magnetic fields has returned our lab to earlier work on magnetic field effects that arise from spin-correlated radical pairs. Understanding the origin(s) of these effects may help to develop these effects as an imaging method.
Model Membranes
Our group has developed supported lipid bilayers as mimics for cell surfaces and tools in biotechnology. A broad vision is to engineer interfaces between hard surfaces and soft materials, ultimately leading to sophisticated biocompatible interfaces that can be used to control, interrogate or organize complex living systems. We have developed methods to partition and manipulate elements of these unique self-assembled systems; these methods are now used in many laboratories.
Recent work addresses three interrelated areas: 1) characterization of membrane organization, domains and protein associations using a novel type of imaging mass spectrometry; 2) models for membrane fusion and investigations into the fusion of enveloped viruses and lipid nanoparticles to their target membrane; 3) development of tethered lipid bilayers as a platform to study membrane domains and junction topology and organization between supported membranes and natural cell membranes.
Energy and Electron Transfer in Photosynthesis
Light-driven long-distance electron transfer in photosynthetic reaction centers is one of the fastest known chemical reactions. We studied this by femtosecond fluorescence and transient absorption spectroscopy, manipulation in electric fields, site-specific and global mutagenesis and some novel types of Stark spectroscopy. While this phase of our work is complete, we continue to use elements of the photosynthetic apparatus to study the electric fields in membranes created in response to transmembrane potential.