The Application of an Emerging Technique forProtein–Protein Interaction Interface Mapping:The Combination of Photo-Initiated Cross-LinkingProtein Nanoprobes with Mass Spectrometry

International Journal of Molecular Sciences

Stiborová Marie¹  Hudeček Jiří¹  Ječmen Tomáš²  Novák Petr²  Šulc Miroslav²  Ptáčková Renata² 

[1] Department of Biochemistry, Faculty of Science, Charles University, Hlavova 2030,CZ-12843 Prague 2, Czech Republic;Institute of Microbiology v.v.i., Academy of Sciences of the Czech Republic,Vídeňská 1083, CZ-14220 Prague 4, Czech Republic;

关键词: 14-3-3ζ homodimer;    protein-protein interaction;    photo cross-linking;    protein nanoprobe;    mass spectrometry;   

DOI  :  10.3390/ijms15069224

来源: DOAJ

【 摘 要 】

Protein–protein interaction was investigated using a protein nanoprobe capable of photo-initiated cross-linking in combination with high-resolution and tandem mass spectrometry. This emerging experimental approach introduces photo-analogs of amino acids within a protein sequence during its recombinant expression, preserves native protein structure and is suitable for mapping the contact between two proteins. The contact surface regions involved in the well-characterized interaction between two molecules of human14-3-3ζ regulatory protein were used as a model. The employed photo-initiated cross-linking techniques extend the number of residues shown to be within interaction distance in the contact surface of the 14-3-3ζ dimer (Gln8–Met78). The results of this study are in agreement with our previously published data from molecular dynamic calculations based on high-resolution chemical cross-linking data and Hydrogen/Deuterium exchange mass spectrometry. The observed contact is also in accord with the 14-3-3ζ X-ray crystal structure (PDB 3dhr). The results of the present work are relevant to the structural biology of transient interaction in the 14-3-3ζ protein, and demonstrate the ability of the chosen methodology (the combination of photo-initiated cross-linking protein nanoprobes and mass spectrometry analysis) to map the protein-protein interface or regions with a flexible structure.

【 授权许可】

Unknown