Tunable paramagnetic relaxation enhancements by [Gd(DPA) 3 ] 3- for protein structure analysis

Abstract

Paramagnetic relaxation enhancements (PRE) present a powerful source of structural information in nuclear magnetic resonance (NMR) studies of proteins and protein-ligand complexes. In contrast to conventional PRE reagents that are covalently attached to the protein, the complex between gadolinium and three dipicolinic acid (DPA) molecules, [Gd(DPA)3]3-, can bind to proteins in a non-covalent yet site-specific manner. This offers straightforward access to PREs that can be scaled by using different ratios of [Gd(DPA)3]3- to protein, allowing quantitative distance measurements for nuclear spins within about 15 Å of the Gd3+ ion. Such data accurately define the metal position relative to the protein, greatly enhancing the interpretation of pseudocontact shifts induced by [Ln(DPA)3]3- complexes of paramagnetic lanthanide (Ln 3+) ions other than gadolinium. As an example we studied the quaternary structure of the homodimeric GCN4 leucine zipper.