Interactions of Substrate and Product Radicals with CoII in Cobalamin and with the Active Site in Ethanolamine Deaminase, Characterized by ESE-EPR and 14N ESEEM Spectroscopies

Abstract

Interactions of the cryotrapped 2-aminopropanol-1-yl substrate radical and aminoethanol-derived product radical catalytic intermediates with the active site of vitamin B12 coenzyme-dependent ethanolamine deaminase from Salmonella typhimurium in disordered frozen aqeuous solution have been characterized by using X-band electron spin−echo electron paramagnetic resonance (ESE-EPR) and electron spin−echo envelope modulation (ESEEM) spectroscopies performed at 6 K. ESE-EPR spectra show that the doublet splitting of the radical line shape by electron−electron exchange and dipolar interactions with CoII in cob(II)alamin is stronger for the substrate radical (11.1 mT) than for the product radical (7.1 mT). The aminoethanol-derived product radical unpaired spin density at C2 is therefore positioned closer to CoII than the 2-aminopropanol-1-yl substrate radical unpaired spin density at C1. Multifrequency three-pulse ESEEM spectra obtained at g = 2.00 for each radical display the same τ- and magnetic field-dependent...