Quantitative, directional measurement of electric field heterogeneity in the active site of ketosteroid isomerase

摘要

Extensive structural studies of enzymes have revealed that biological catalysis occurs within sequestered active site crevices, where reacting substrates are surrounded by an anisotropic and chemically complex constellation of charged, polar, and hydrophobic groups. However, translating these structural pictures into a rigorous, quantitative understanding of the highly idiosyncratic electrostatic environment within a protein interior has remained one of the most challenging and elusive goals of protein biophysics. To address the scarcity of incisive experimental measures of electrostatic properties within proteins we have developed the nitrile (carbon-nitrogen triple bond) as a spectroscopic probe capable of reporting on local electrostatic fields.