title: Information transfer in the penta-EF-hand protein sorcin does not operate via the canonical structural/functional pairing. A study with site-specific mutants.
abstract: Sorcin is a typical penta-EF-hand protein that participates in Ca2+-regulated processes by translocating reversibly from cytosol to membranes, where it interacts with different target proteins in different tissues. Binding of two Ca2+/monomer triggers translocation, although EF1, EF2, and EF3 are potentially able to bind calcium at micromolar concentrations. To identify the functional pair, the conserved bidentate -Z glutamate in these EF-hands was mutated to yield E53Q-, E94A-, and E124A-sorcin, respectively. Limited structural perturbations occur only in E124A-sorcin due to involvement of Glu-124 in a network of interactions that comprise the long D helix connecting EF3 to EF2. The overall affinity for Ca2+ and for two sorcin targets, annexin VII and the ryanodine receptor, follows the order wild-type > E53Q- > E94A- > E124A-sorcin, indicating that disruption of EF3 has the largest functional impact and that disruption of EF2 and EF1 has progressively smaller effects. Based on this experimental evidence, EF3 and EF2, which are not paired in the canonical manner, are the functional EF-hands. Sorcin is proposed to be activated upon Ca2+ binding to EF3 and transmission of the conformational change at Glu-124 via the D helix to EF2 and from there to EF1 via the canonical structural/functional pairing. This mechanism may be applicable to all penta-EF-hand proteins.
authors.affiliation: Consiglio Nazionale delle Ricerche Institute of Molecular Biology and Pathology, Department of Biochemical Sciences A. Rossi Fanelli, University of Rome La Sapienza, Italy.
authors.forename: Manuela
authors.initials: M
authors.lastname: Mella
authors.forename: Gianni
authors.initials: G
authors.lastname: Colotti
authors.forename: Carlotta
authors.initials: C
authors.lastname: Zamparelli
authors.forename: Daniela
authors.initials: D
authors.lastname: Verzili
authors.forename: Andrea
authors.initials: A
authors.lastname: Ilari
authors.forename: Emilia
authors.initials: E
authors.lastname: Chiancone
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D002942:Circular Dichroism; D006224:Cricetinae; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D013329:Structure-Activity Relationship
chemical_list: D002135:Calcium-Binding Proteins; D002118:Calcium