Investigating the intermediates in the reaction of ribonucleoside triphosphate reductase from Lactobacillus leichmannii: An application of HF EPR-RFQ technology |
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Authors: | Manzerova Julia Krymov Vladimir Gerfen Gary J |
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Institution: | Department of Physiology and Biophysics, Albert Einstein College of Medicine, 1300 Morris Park Ave., Bronx, NY 10461, United States. |
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Abstract: | In this investigation high-frequency electron paramagnetic resonance spectroscopy (HFEPR) in conjunction with innovative rapid freeze-quench (RFQ) technology is employed to study the exchange-coupled thiyl radical-cob(II)alamin system in ribonucleotide reductase from a prokaryote Lactobacillus leichmannii. The size of the exchange coupling (Jex) and the values of the thiyl radical g tensor are refined, while confirming the previously determined (Gerfen et al. (1996) 20]) distance between the paramagnets. Conclusions relevant to ribonucleotide reductase catalysis and the architecture of the active site are presented. A key part of this work has been the development of a unique RFQ apparatus for the preparation of millisecond quench time RFQ samples which can be packed into small (0.5 mm ID) sample tubes used for CW and pulsed HFEPR--lack of this ability has heretofore precluded such studies. The technology is compatible with a broad range of spectroscopic techniques and can be readily adopted by other laboratories. |
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