Abstract
Arginine side-chains play a distinct role because of their high pKa and perpetual positive charge. An NMR method is presented, based on carbon-detected 13Cζ–15Ne correlation spectra, which allows probing the arginine side-chains and their dynamics at neutral-to-high pH. The methodology is demonstrated on human histone deacetylase 8.
Highlights
Arginine has a unique role among the 20 standard proteinogenic amino acids
Relying on detection of aliphatic protons. Whilst this method circumvents the rapid exchange and allows measurements of the 15Ne chemical shift of arginine, it is less beneficial for nuclear spin relaxation measurements and for studying larger proteins, where the two bound protons enhance the transverse relaxation of the aliphatic carbons
Magnetization is transfered from 13Cz to 15Ne via the scalar coupling, while transfers from 13Cz to 15Nh are avoided by applying a selective 15Ne inversion pulse in the INEPT transfer steps
Summary
Arginine has a unique role among the 20 standard proteinogenic amino acids. In contrast to other charged amino acid side-chains the charge of arginine side-chains is relatively unaffected by the surrounding environment.[1]. We describe an NMR pulse scheme for probing arginine side-chains and their dynamics at neutral-to-high pH by carbon-detected 13Cz–15Ne correlated spectra (Figure 1). As a first application we probed the arginine side-chains of T4 lysozyme L99A (T4L L99A; 18 kDa) shown in Figure 2 a.
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