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J Mol Biol. 2019 Jul 12;431(15):2869-2883. doi: 10.1016/j.jmb.2019.05.008. Epub 2019 May 11.

SARAF Luminal Domain Structure Reveals a Novel Domain-Swapped β-Sandwich Fold Important for SOCE Modulation.

Author information

1
Cardiovascular Research Institute, University of California, San Francisco, CA 94148, USA.
2
Department of Biomolecular Sciences, Weizmann Institute of Science, Rehovot 7610001, Israel.
3
Rappaport Family School of Medicine, Technion-Israel Institute of Technology, Haifa 31096, Israel.
4
Department of Biomolecular Sciences, Weizmann Institute of Science, Rehovot 7610001, Israel. Electronic address: e.reuveny@weizmann.ac.il.
5
Cardiovascular Research Institute, University of California, San Francisco, CA 94148, USA; Department of Biochemistry and Biophysics, University of California, San Francisco, CA 94158, USA; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, CA 94158, USA; California Institute for Quantitative Biomedical Research, University of California, San Francisco, CA 94158, USA; Kavli Institute for Fundamental Neuroscience, University of California, San Francisco, San Fancisco, CA 94158, USA; Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA. Electronic address: daniel.minor@ucsf.edu.

Abstract

Store-Operated Calcium Entry (SOCE) plays key roles in cell proliferation, muscle contraction, immune responses, and memory formation. The coordinated interactions of a number of proteins from the plasma and endoplasmic reticulum membranes control SOCE to replenish internal Ca2+ stores and generate intracellular Ca2+ signals. SARAF, an endoplasmic reticulum resident component of the SOCE pathway having no homology to any characterized protein, serves as an important brake on SOCE. Here, we describe the X-ray crystal structure of the SARAF luminal domain, SARAFL. This domain forms a novel 10-stranded β-sandwich fold that includes a set of three conserved disulfide bonds, denoted the "SARAF-fold." The structure reveals a domain-swapped dimer in which the last two β-strands (β9 and β10) are exchanged forming a region denoted the "SARAF luminal switch" that is essential for dimerization. Sequence comparisons reveal that the SARAF-fold is highly conserved in vertebrates and in a variety of pathologic fungi. Förster resonance energy transfer experiments using full-length SARAF validate the formation of the domain-swapped dimer in cells and demonstrate that dimerization is reversible. A designed variant lacking the SARAF luminal switch shows that the domain swapping is essential to function and indicates that the SARAF dimer accelerates SOCE inactivation.

KEYWORDS:

SARAF; X‐ray crystallography; domain swapping; electrophysiology; store-operated calcium entry (SOCE); β-sandwich fold

PMID:
31082439
PMCID:
PMC6599547
[Available on 2020-07-12]
DOI:
10.1016/j.jmb.2019.05.008

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