Chapter 6
PDZ Sample Quality Assessment by Biochemical
and Biophysical Characterizations
Ce ´ lia Caillet-Saguy, Se ´ bastien Bru ˆ le ´ , Nicolas Wolff, and Bertrand Raynal
Abstract
PDZ domains are small globular domains involved in protein–protein interactions. They participate in a
wide range of critical cellular processes. These domains, very abundant in the human proteome, are widely
studied by high-throughput interactomics approaches and by biophysical and structural methods. However, the quality of the results is strongly related to the optimal folding and solubility of the domains. We
provide here a detailed description of protocols for a strict quality assessment of the PDZ constructs. We
describe appropriate experimental approaches that have been selected to overcome the small size of such
domains to check the purity, identity, homogeneity, stability, and folding of samples.
Key words PDZ domain, Folding, Protein sample, Quality control, Biophysical techniques, Stability,
Purity, Identity, Structure, Quantification
1 Introduction
PDZ domains are one of the most abundant protein–protein interaction domains found in metazoans. The human proteome contains 266 identified PDZ domains, termed the PDZome, spread
over 152 proteins [1]. PDZ domains are mainly found in large
scaffold proteins, existing with a wide variety of modular domains.
The PDZ-mediated interactions participate in numerous critical
cellular processes and are often targeted in pathologies and by
numerous viral proteins during infection [2].
PDZs are globular domains of ~90 amino-acid residues, which
recognize PDZ binding motifs (PBMs), mainly at the extreme
C-terminus of their partner proteins. They are recognized as wellfolded and compact domains. The boundaries of most experimental
PDZ constructs that have been used for structural studies or interaction assays generally extend beyond the strict boundaries of the
Jean-Paul Borg (ed.), PDZ Mediated Interactions: Methods and Protocols, Methods in Molecular Biology, vol. 2256,
https://doi.org/10.1007/978-1-0716-1166-1_6, © Springer Science+Business Media, LLC, part of Springer Nature 2021
Ce ´lia Caillet-Saguy and Se ´bastien Bru ˆle ´ contributed equally to this work.
89
PDZ Sample Quality Assessment by Biochemical
and Biophysical Characterizations
Ce ´ lia Caillet-Saguy, Se ´ bastien Bru ˆ le ´ , Nicolas Wolff, and Bertrand Raynal
Abstract
PDZ domains are small globular domains involved in protein–protein interactions. They participate in a
wide range of critical cellular processes. These domains, very abundant in the human proteome, are widely
studied by high-throughput interactomics approaches and by biophysical and structural methods. However, the quality of the results is strongly related to the optimal folding and solubility of the domains. We
provide here a detailed description of protocols for a strict quality assessment of the PDZ constructs. We
describe appropriate experimental approaches that have been selected to overcome the small size of such
domains to check the purity, identity, homogeneity, stability, and folding of samples.
Key words PDZ domain, Folding, Protein sample, Quality control, Biophysical techniques, Stability,
Purity, Identity, Structure, Quantification
1 Introduction
PDZ domains are one of the most abundant protein–protein interaction domains found in metazoans. The human proteome contains 266 identified PDZ domains, termed the PDZome, spread
over 152 proteins [1]. PDZ domains are mainly found in large
scaffold proteins, existing with a wide variety of modular domains.
The PDZ-mediated interactions participate in numerous critical
cellular processes and are often targeted in pathologies and by
numerous viral proteins during infection [2].
PDZs are globular domains of ~90 amino-acid residues, which
recognize PDZ binding motifs (PBMs), mainly at the extreme
C-terminus of their partner proteins. They are recognized as wellfolded and compact domains. The boundaries of most experimental
PDZ constructs that have been used for structural studies or interaction assays generally extend beyond the strict boundaries of the
Jean-Paul Borg (ed.), PDZ Mediated Interactions: Methods and Protocols, Methods in Molecular Biology, vol. 2256,
https://doi.org/10.1007/978-1-0716-1166-1_6, © Springer Science+Business Media, LLC, part of Springer Nature 2021
Ce ´lia Caillet-Saguy and Se ´bastien Bru ˆle ´ contributed equally to this work.
89