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Old 11-09-2017, 08:55 AM
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Default Proteinâ??nucleotide contacts in motor proteins detected by DNP-enhanced solid-state NMR

Proteinâ??nucleotide contacts in motor proteins detected by DNP-enhanced solid-state NMR

Abstract

DNP (dynamic nuclear polarization)-enhanced solid-state NMR is employed to directly detect proteinâ??DNA and proteinâ??ATP interactions and identify the residue type establishing the intermolecular contacts. While conventional solid-state NMR can detect proteinâ??DNA interactions in large oligomeric protein assemblies in favorable cases, it typically suffers from low signal-to-noise ratios. We show here, for the oligomeric DnaB helicase from Helicobacter pylori complexed with ADP and single-stranded DNA, that this limitation can be overcome by using DNP-enhanced spectroscopy. Interactions are established by DNP-enhanced 31Pâ??13C polarization-transfer experiments followed by the recording of a 2D 13Câ??13C correlation experiment. The NMR spectra were obtained in less than 2 days and allowed the identification of residues of the motor protein involved in nucleotide binding.



Source: Journal of Biomolecular NMR
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