Journal article
Isotopic Labeling of Formate Dehydrogenase Perturbs the Protein Dynamics
The journal of physical chemistry. B, Vol.123(49), pp.10403-10409
12/12/2019
DOI: 10.1021/acs.jpcb.9b08426
PMID: 31696711
Abstract
Isotope substitution of enzymes has become a means of addressing the participation of protein motions in enzyme-catalyzed reactions. The idea is that only the enzyme mass will be altered and not the electrostatics, so that the protein dynamics are essentially the same but at lower frequencies because of the mass change. In this study, we variably label all carbon atoms in formate dehydrogenase (FDH) with 13C, all nitrogen atoms with 15N, and all nonexchangeable hydrogen atoms with deuterium and investigate the impact that isotopic substitution has on the dynamics at the active site by two-dimensional infrared spectroscopy and compare with the measurements of the temperature dependence of the intrinsic kinetic isotope effects (KIEs). We show that 15N labeling of FDH has the largest effect and makes the active site more heterogeneous, whereas the addition of nonexchangeable deuterium appears to have the opposite effect of 15N on active-site dynamics, resulting in a behavior similar to that of native FDH. Nevertheless, the temperature dependence of the KIEs shows a monotonic trend with protein mass that does not correspond with the changes in dynamics. These results suggest that isotope labeling has more than just a mass effect on enzyme dynamics and may influence electrostatics in ways that complicate the interpretation of the protein isotope effect.
Details
- Title: Subtitle
- Isotopic Labeling of Formate Dehydrogenase Perturbs the Protein Dynamics
- Creators
- Chethya Ranasinghe - Department of ChemistryPhilip Pagano - Department of ChemistryPaul J Sapienza - Division of Chemical Biology and Medicinal Chemistry, UNC Eshelman School of PharmacyAndrew L Lee - Division of Chemical Biology and Medicinal Chemistry, UNC Eshelman School of PharmacyAmnon Kohen - Department of ChemistryChristopher M Cheatum - Department of Chemistry
- Resource Type
- Journal article
- Publication Details
- The journal of physical chemistry. B, Vol.123(49), pp.10403-10409
- DOI
- 10.1021/acs.jpcb.9b08426
- PMID
- 31696711
- NLM abbreviation
- J Phys Chem B
- ISSN
- 1520-6106
- eISSN
- 1520-5207
- Publisher
- American Chemical Society
- Grant note
- DOI: 10.13039/100000001, name: National Science Foundation, award: CHE-1707598
- Language
- English
- Date published
- 12/12/2019
- Academic Unit
- Liberal Arts and Science Admin; Chemistry
- Record Identifier
- 9984216599702771
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