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Molecular Basis of 14‐3‐3 Protein Dependent Regulation of Caspase‐2
Author(s) -
Obsilova Veronika,
Kalabova Dana,
Filandr Frantisek,
Alblova Miroslava,
Petrvalska Olivia,
Horvath Matej,
Man Petr,
Obsil Tomas
Publication year - 2020
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2020.34.s1.02861
Subject(s) - chemistry , signal transducing adaptor protein , microbiology and biotechnology , linker , phosphorylation , cleavage (geology) , gene isoform , biophysics , biochemistry , biology , paleontology , fracture (geology) , computer science , gene , operating system
Among all species, caspase‐2 (C2) is the most evolutionarily conserved caspase required for effective initiation of apoptosis following death stimuli. C2 is activated through dimerization and autoproteolytic cleavage and inhibited through phosphorylation at Ser 139 and Ser 164 , within the linker between the caspase recruitment and p19 domains of the zymogen, followed by association with the adaptor protein 14‐3‐3, which maintains C2 in its immature form procaspase (proC2). However, the mechanism of 14‐3‐3‐dependent inhibition of C2 activation remains unclear. Here, we report the structural characterization of the complex between proC2 and 14‐3‐3 by hydrogen/deuterium mass spectrometry (HDX‐MS) and protein crystallography to determine the molecular basis for 14‐3‐3‐mediated inhibition of C2 activation. Our data reveal that the 14‐3‐3 dimer interacts with proC2 not only through ligand‐binding grooves but also through other regions outside the central channel, thus explaining the isoform‐dependent specificity of 14‐3‐3 protein binding to proC2 and the substantially higher binding affinity of 14‐3‐3 protein to proC2 than to the doubly phosphorylated peptide. The formation of the complex between 14‐3‐3 protein and proC2 does not induce any large conformational change in proC2. Furthermore, 14‐3‐3 protein interacts with and masks both the nuclear localization sequence (NLS) and the C‐terminal region of the p12 domain of proC2 through transient interactions in which both the p19 and p12 domains of proC2 are not firmly docked onto the surface of 14‐3‐3. This masked region of p12 domain is involved in caspase‐2 dimerization. Therefore, 14‐3‐3 protein likely inhibits proC2 activation by blocking its dimerization surface. Support or Funding Information Funding This study was funded by the Czech Science Foundation (V.O., grant number 19‐00121S), the Czech Academy of Sciences (RVO:67985823 of the Institute of Physiology), project BIOCEV (CZ.1.05/1.1.00.02.0109 and LQ1604) and MEYS CR (LM2015043 CIISB, Biocev, Biophysical methods, Structural Mass Spectrometry).Crystal structure of the caspase‐2 peptide pS139+pS164 bound to the 14‐3‐3 dimer.