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Site-specific tagging proteins with a rigid, small and stable transition metal chelator, 8-hydroxyquinoline, for paramagnetic NMR analysis

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Abstract

Design of a paramagnetic metal binding motif in a protein is a valuable way for understanding the function, dynamics and interactions of a protein by paramagnetic NMR spectroscopy. Several strategies have been proposed to site-specifically tag proteins with paramagnetic lanthanide ions. Here we report a simple approach of engineering a transition metal binding motif via site-specific labelling of a protein with 2-vinyl-8-hydroxyquinoline (2V-8HQ). The protein-2V-8HQ adduct forms a stable complex with transition metal ions, Mn(II), Co(II), Ni(II), Cu(II) and Zn(II). The paramagnetic effects generated by these transition metal ions were evaluated by NMR spectroscopy. We show that 2V-8HQ is a rigid and stable transition metal binding tag. The coordination of the metal ion can be assisted by protein sidechains. More importantly, tunable paramagnetic tensors are simply obtained in an α-helix that possesses solvent exposed residues in positions i and i + 3, where i is the residue to be mutated to cysteine, i + 3 is Gln or Glu or i − 4 is His. The coordination of a sidechain carboxylate/amide or imidazole to cobalt(II) results in different structural geometries, leading to different paramagnetic tensors as shown by experimental data.

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Acknowledgments

Financial support by the 973 program (2013CB910200), the National Science Foundation of China (21473095 and 21273121), and the Australian Research Council (DP120100561 and DP150100383) is greatly acknowledged.

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Authors and Affiliations

  1. State Key Laboratory of Elemento-Organic Chemistry, Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Nankai University, Tianjin, 300071, China

    Yin Yang, Feng Huang & Xun-Cheng Su

  2. Research School of Chemistry, Australian National University, Canberra, ACT, 0200, Australia

    Thomas Huber

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  1. Yin Yang
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  2. Feng Huang
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  4. Xun-Cheng Su
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Correspondence to Xun-Cheng Su.

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Yang, Y., Huang, F., Huber, T. et al. Site-specific tagging proteins with a rigid, small and stable transition metal chelator, 8-hydroxyquinoline, for paramagnetic NMR analysis. J Biomol NMR 64, 103–113 (2016). https://doi.org/10.1007/s10858-016-0011-7

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