1. Rubredoxin as a paramagnetic relaxation-inducing probe
- Author
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Almeida, Rui M., Pauleta, Sofia R., Moura, Isabel, and Moura, José J.G.
- Subjects
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IRON proteins , *PARAMAGNETISM , *CHARGE exchange , *NUCLEAR magnetic resonance spectroscopy , *CYTOCHROME c , *HEME , *SIMULATION methods & models , *RELAXATION phenomena - Abstract
Abstract: The paramagnetic effect due to the presence of a metal center with unpaired electrons is no longer considered a hindrance in protein NMR spectroscopy. In the present work, the paramagnetic effect due to the presence of a metal center with unpaired electrons was used to map the interface of an electron transfer complex. Desulfovibrio gigas cytochrome c 3 was chosen as target to study the effect of the paramagnetic probe, Fe-rubredoxin, which produced specific line broadening in the heme IV methyl resonances M21 and M181. The rubredoxin binding surface in the complex with cytochrome c 3 was identified in a heteronuclear 2D NMR titration. The identified heme methyls on cytochrome c 3 are involved in the binding interface of the complex, a result that is in agreement with the predicted complexes obtained by restrained molecular docking, which shows a cluster of possible solutions near heme IV. The use of a paramagnetic probe in 1HNMR titration and the mapping of the complex interface, in combination with a molecular simulation algorithm proved to be a valuable strategy to study electron transfer complexes involving non-heme iron proteins and cytochromes. [Copyright &y& Elsevier]
- Published
- 2009
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