Engineering [Ln(DPA)3]3â?? binding sites in proteins: a widely applicable method for tagging proteins with lanthanide ions

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Engineering [Ln(DPA)3]3â?? binding sites in proteins: a widely applicable method for tagging proteins with lanthanide ions


Abstract Paramagnetic relaxation enhancements from unpaired electrons observed in nuclear magnetic resonance (NMR) spectra present powerful long-range distance restraints. The most frequently used paramagnetic tags, however, are tethered to the protein via disulfide bonds, requiring proteins with single cysteine residues for covalent attachment. Here we present a straightforward strategy to tag proteins site-specifically with paramagnetic lanthanides without a tether and independent of cysteine residues. It relies on preferential binding of the complex between three dipicolinic acid molecules (DPA) and a lanthanide ion (Ln3+), [Ln(DPA)3]3â??, to a pair of positively charged amino acids whose charges are not compensated by negatively charged residues nearby. This situation rarely occurs in wild-type proteins, allowing the creation of specific binding sites simply by introduction of positively charged residues that are positioned far from glutamate or aspartate residues. The concept is demonstrated with the hnRNPLL RRM1 domain. In addition, we show that histidine- and arginine-tags present binding sites for [Ln(DPA)3]3â??.

• Content Type Journal Article
• Pages 1-10
• DOI 10.1007/s10858-011-9529-x
• Authors
  • Xinying Jia, Research School of Chemistry, Australian National University, Canberra, ACT 0200, Australia
  • Hiromasa Yagi, Research School of Chemistry, Australian National University, Canberra, ACT 0200, Australia
  • Xun-Cheng Su, Research School of Chemistry, Australian National University, Canberra, ACT 0200, Australia
  • Mitchell Stanton-Cook, Research School of Chemistry, Australian National University, Canberra, ACT 0200, Australia
  • Thomas Huber, Research School of Chemistry, Australian National University, Canberra, ACT 0200, Australia
  • Gottfried Otting, Research School of Chemistry, Australian National University, Canberra, ACT 0200, Australia

• • Journal Journal of Biomolecular NMR
  • Online ISSN 1573-5001
  • Print ISSN 0925-2738

Source: Journal of Biomolecular NMR