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“CON-CON” assignment strategy for highly flexible intrinsically disordered proteins

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Abstract

Intrinsically disordered proteins (IDPs) are a class of highly flexible proteins whose characterization by NMR spectroscopy is complicated by severe spectral overlaps. The development of experiments designed to facilitate the sequence-specific assignment procedure is thus very important to improve the tools for the characterization of IDPs and thus to be able to focus on IDPs of increasing size and complexity. Here, we present and describe the implementation of a set of novel 1H-detected 5D experiments, (HACA)CON(CACO)NCO(CA)HA, BT-(H)NCO(CAN)CONNH and BT-HN(COCAN)CONNH, optimized for the study of highly flexible IDPs that exploit the best resolved correlations, those involving the carbonyl and nitrogen nuclei of neighboring amino acids, to achieve sequence-specific resonance assignment. Together with the analogous recently proposed pulse schemes based on 13C detection, they form a complete set of experiments for sequence-specific assignment of highly flexible IDPs. Depending on the particular sample conditions (concentration, lifetime, pH, temperature, etc.), these experiments present certain advantages and disadvantages that will be discussed. Needless to say, that the availability of a variety of complementary experiments will be important for accurate determination of resonance frequencies in complex IDPs.

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Acknowledgments

This work has been supported in part by the European Commission Projects IDPbyNMR (Contract No. 264257), BioNMR (Contract No. 261863) and INSTRUCT (Contract No. 211252).

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Authors and Affiliations

  1. CERM and Department of Chemistry, University of Florence, Via Luigi Sacconi 6, 50019, Sesto Fiorentino, Florence, Italy

    Alessandro Piai, Tomáš Hošek, Leonardo Gonnelli, Roberta Pierattelli & Isabella C. Felli

  2. Faculty of Chemistry, Biological and Chemical Research Centre, University of Warsaw, Żwirki i Wigury 101, 02-089, Warsaw, Poland

    Anna Zawadzka-Kazimierczuk & Wiktor Koźmiński

  3. Institut de Biologie Structurale, Université Grenoble 1, CNRS, CEA, Rue Jules Horowitz 41, 38027, Grenoble Cedex 1, France

    Bernhard Brutscher

  4. Bruker BioSpin GmbH, Silberstreifen, 76287, Rheinstetten, Germany

    Wolfgang Bermel

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  1. Alessandro Piai
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  2. Tomáš Hošek
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  3. Leonardo Gonnelli
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Correspondence to Isabella C. Felli.

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Piai, A., Hošek, T., Gonnelli, L. et al. “CON-CON” assignment strategy for highly flexible intrinsically disordered proteins. J Biomol NMR 60, 209–218 (2014). https://doi.org/10.1007/s10858-014-9867-6

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