The 14-3-3/SLP76 protein-protein interaction in T-cell receptor signalling: a structural and biophysical characterization

Lorenzo Soini; Seppe Leysen; Jeremy Davis; Marta Westwood; Christian Ottmann

doi:10.1002/1873-3468.13993

The 14-3-3/SLP76 protein-protein interaction in T-cell receptor signalling: a structural and biophysical characterization

FEBS Lett. 2021 Feb;595(3):404-414. doi: 10.1002/1873-3468.13993. Epub 2020 Nov 22.

Authors

Lorenzo Soini^{1

2}, Seppe Leysen³, Jeremy Davis², Marta Westwood⁴, Christian Ottmann¹

Affiliations

¹ Laboratory of Chemical Biology, Department of Biomedical Engineering and Institute for Complex Molecular Systems, Eindhoven University of Technology, Eindhoven, the Netherlands.
² Department of Chemistry, UCB Celltech, Slough, UK.
³ Department of Structural Biology and Biophysics, UCB Celltech, Slough, UK.
⁴ Discovery Science, UCB Celltech, Slough, UK.

PMID: 33159816
DOI: 10.1002/1873-3468.13993

Abstract

The SH2 domain-containing protein of 76 kDa, SLP76, is an important adaptor protein that coordinates a complex protein network downstream of T-cell receptors (TCR), ultimately regulating the immune response. Upon phosphorylation on Ser376, SLP76 interacts with 14-3-3 adaptor proteins, which leads to its proteolytic degradation. This provides a negative feedback mechanism by which TCR signalling can be controlled. To gain insight into the 14-3-3/SLP76 protein-protein interaction (PPI), we have determined a high-resolution crystal structure of a SLP76 synthetic peptide containing Ser376 with 14-3-3σ. We then characterized its binding to 14-3-3 proteins biophysically by means of fluorescence polarization and isothermal titration calorimetry. Furthermore, we generated two recombinant SLP76 protein constructs and characterized their binding to 14-3-3. Our work lays the foundation for drug design efforts aimed at targeting the 14-3-3/SLP76 interaction and, thereby, TCR signalling.

Keywords: TCR signalling and 14-3-3; X-ray protein crystallography; isothermal titration calorimetry; protein constructs; surface plasmon resonance.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

14-3-3 Proteins / chemistry*
14-3-3 Proteins / genetics
14-3-3 Proteins / immunology
Adaptor Proteins, Signal Transducing / chemistry*
Adaptor Proteins, Signal Transducing / genetics
Adaptor Proteins, Signal Transducing / immunology
Binding Sites
Cloning, Molecular
Crystallography, X-Ray
Escherichia coli / genetics
Escherichia coli / metabolism
Feedback, Physiological
Gene Expression
Gene Expression Regulation
Genetic Vectors / chemistry
Genetic Vectors / metabolism
Humans
Immunity, Innate
Kinetics
Models, Molecular
Mutation
Peptides / chemistry*
Peptides / genetics
Peptides / immunology
Phosphoproteins / chemistry*
Phosphoproteins / genetics
Phosphoproteins / immunology
Phosphorylation
Protein Binding
Protein Conformation, alpha-Helical
Protein Conformation, beta-Strand
Protein Interaction Domains and Motifs
Receptors, Antigen, T-Cell / genetics
Receptors, Antigen, T-Cell / immunology
Recombinant Proteins / chemistry
Recombinant Proteins / genetics
Recombinant Proteins / metabolism
Signal Transduction / genetics*
Signal Transduction / immunology
T-Lymphocytes / cytology
T-Lymphocytes / immunology

Substances

14-3-3 Proteins
Adaptor Proteins, Signal Transducing
Peptides
Phosphoproteins
Receptors, Antigen, T-Cell
Recombinant Proteins
SLP-76 signal Transducing adaptor proteins
YWHAG protein, human
YWHAZ protein, human