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SH2 Domains pp 269-290 | Cite as

NMR Chemical Shift Mapping of SH2 Peptide Interactions

  • Marissa A. McKercher
  • Deborah S. Wuttke
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1555)

Abstract

Heteronuclear single quantum coherence (HSQC) nuclear magnetic resonance (NMR) experiments offer a rapid and high resolution approach to gaining binding and conformational insights into a protein–peptide interaction. By tracking 1H and 15N chemical shift changes over the course of a peptide titration into isotopically labeled protein, amide NH pairs of amino acids whose chemical environment changes upon peptide binding can be identified. When mapped onto a structure of the protein, this approach can identify the peptide-binding interface or regions undergoing conformation changes within a protein upon ligand binding. Monitoring NMR chemical shift changes can also serve as a screening technique to identify novel interaction partners for a protein or to determine the binding affinity of a weak protein–peptide interaction. Here, we describe the application of NMR chemical shift mapping to the study of peptide binding to the C-terminal SH2 domain of PLCγ1.

Key words

Nuclear magnetic resonance (NMR) SH2 domain Peptide Chemical shift mapping (CSM) Phosphotyrosine PLCγ1 

Notes

Acknowledgments

We would like to thank our collaborators Dr. Zhongping Tan and Dr. Xiaoyang Guan for the phosphopeptide synthesis, as well as Dr. Julie Forman-Kay for kindly providing us with a PLCγ1-pET11d construct. We would also like to thank Sabrina Hunt and Kathryn Wall for their generous feedback. This work was supported by NSF grant MCB1121842 (to D.S.W.).

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Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  1. 1.Department of Chemistry and BiochemistryUniversity of ColoradoBoulderUSA

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