Electrical perturbation of solvent dynamics using open-ended coaxial probe Open Access

Abstract

Enzyme function is intimately related to motions in the surrounding solvent environment. Ethanolamine ammonia-lyase from Salmonella typhimurium exemplifies an enzyme in which a select class of solvent-coupled protein configurational fluctuations is obligatory for physiological activity. The ability to drive specific classes of solvent-coupled protein fluctuations by using an electric field oscillating at microwave frequencies provides a mechanism for actuating and characterizing the solvent-protein dynamical coupling. As a first step, we design a microwave circuit and test system to probe the dynamics in aqueous solution, by using an open-ended coaxial probe to deliver microwave radiation to interstitial solvent domains in frozen solution samples (200 - 240 K) in the frequency range of 1.0 - 1.6 gigahertz, with a maximum power of 2 Watts. Continuous-wave electron paramagnetic resonance (EPR) spectroscopy is used for reporting on the solvent dynamics through the rotational mobility of the nitroxide spin probe, TEMPOL. We design a circuit featuring a circulator component, which allows for delivery of microwave power to the sample via the coaxial probe, while reporting on the power absorption by the sample. A microwave power- and frequency-dependent increase in amplitude and associated narrowing of the EPR spectra is observed, which indicates increased mobility in the solvent. The devised twin capabilities of microwave actuation and sensitive detection of solvent motion signify the ability to characterize the role of select, gigahertz-frequency fluctuations in enzyme catalysis.

Table of Contents

Introduction     1

Microwave effects on enzyme activity     1 Ethanolamine ammonia-lyase as an example system     2 Open-ended Coaxial probe technique     2 Electron paramagnetic resonance (EPR) spectroscopy     5 Circuit Design     7 Motivation     9

Methods    10

Sample preparation for dielectric property change measurements    10 EPR Sample Preparation    10 Cooling bath and temperature control experiments    10 Continuous wave EPR spectroscopy    11 Application of probe microwave power    11

Results    12

Detection of differences in dielectric properties with open-ended coaxial probe       12 Actuation of solvent mobility in CW EPR experiment    14 Power dependence of solvent dynamical effect    16 Frequency dependence of solvent dynamical effect    18

Discussion

Confirmation of appropriate microwave power absorption dependence on material dielectric properties    19 Subtraction analysis of 235 K EPR spectra reveals effect of microwave irradiation on spin probe rotational motion    20 Verification of line shape narrowing as an indicator of increased solvent mobility    23 Conclusion     25

References    26

About this Honors Thesis

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School	Emory College
Department	Physics
Degree	B.S.
Submission	Honors Thesis
Language	English
Research Field	Biophysics, General Physics, Molecular Physics, Electricity and Magnetism
Keyword	Solvent dynamics Microwave Electrical perturbation EPR Spin probe
Committee Chair / Thesis Advisor	Dr. Kurt Warncke, Emory University
Committee Members	Dr. Wladimir Benalcazar, Emory University Dr. Jed Brody, Emory University

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