The structure of Ca2+sensor case16 reveals the mechanism of reaction to low Ca2+concentrations

Lukas Leder, Wilhelm Stark, Felix Freuler, May Marsh, Marco Meyerhofer, Thomas Stettler, Lorenz M. Mayr, Olga V. Britanova, Lydia A. Strukova, Dmitriy M. Chudakov, Ekaterina A. Souslova

Research output: Contribution to journalArticlepeer-review

14 Citations (Scopus)

Abstract

Here we report the first crystal structure of a high-contrast genetically encoded circularly permuted green fluorescent protein (cpGFP)-based CaCa2+sensor, Case16, in the presence of a low CaCa2+concentration. The structure reveals the positioning of the chromophore within Case16 at the first stage of the Ca2+-dependent response when only two out of four Ca2+-binding pockets of calmodulin (CaM) are occupied with CaCa2+ions. In such a "half Ca2+-bound state", Case16 is characterized by an incomplete interaction between its CaM-/M13-domains. We also report the crystal structure of the related CaCa2+sensor Case12 at saturating Ca Ca2+concentration. Based on this structure, we postulate that cpGFP-based CaCa2+sensors can form non-functional homodimers where the CaM-domainof one sensor molecule binds symmetrically to the M13-peptide of the partner sensor molecule. Case12 and Case16 behavior upon addition of high concentrations of free CaM or M13-peptide reveals that the latter effectively blocks the fluorescent response of the sensor. We speculate that the demonstrated intermolecular interaction with endogenous substrates and homodimerization can impede proper functioning of this type of CaCa2+sensors in living cells.

Original languageEnglish
Pages (from-to)8143-8160
Number of pages18
JournalSensors (Switzerland)
Volume10
Issue number9
DOIs
Publication statusPublished - Sep 2010
Externally publishedYes

Keywords

  • Calcium sensor
  • Circularly permuted green fluorescent protein
  • Crystal structure
  • Fluorescent calcium indicator protein
  • Genetically encoded

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