Tracking Conformational Changes in Calmodulin in vitro, in Cell Extract, and in Cells by Electron Paramagnetic Resonance Distance Measurements

Arina Dalaloyan, Andrea Martorana, Yoav Barak, Diana Gataulin, Eitan Reuveny, Andrew Howe, Michael Elbaum, Shira Albeck, Tamar Unger, Veronica Frydman, Elwy H. Abdelkader, Gottfried Otting, Daniella Goldfarb

Research output: Contribution to journalArticlepeer-review

Abstract

It is an open question whether the conformations of proteins sampled in dilute solutions are the same as in the cellular environment. Here we address this question by double electron-electron resonance (DEER) distance measurements with Gd(III) spin labels to probe the conformations of calmodulin (CaM) in vitro, in cell extract, and in human HeLa cells. Using the CaM mutants N53C/T110C and T34C/T117C labeled with maleimide-DOTA-Gd(III) in the N- and C-terminal domains, we observed broad and varied interdomain distance distributions. The in vitro distance distributions of apo-CaM and holo-CaM in the presence and absence of the IQ target peptide can be described by combinations of closed, open, and collapsed conformations. In cell extract, apo- and holo-CaM bind to target proteins in a similar way as apo- and holo-CaM bind to IQ peptide in vitro. In HeLa cells, however, in the presence or absence of elevated in-cell Ca2+ levels CaM unexpectedly produced more open conformations and very broad distance distributions indicative of many different interactions with in-cell components. These results show-case the importance of in-cell analyses of protein structures.

Original languageEnglish
Pages (from-to)1860-1868
Number of pages9
JournalChemPhysChem
Volume20
Issue number14
Early online date4 May 2019
DOIs
StatePublished - 16 Jul 2019

All Science Journal Classification (ASJC) codes

  • Atomic and Molecular Physics, and Optics
  • Physical and Theoretical Chemistry

Fingerprint

Dive into the research topics of 'Tracking Conformational Changes in Calmodulin in vitro, in Cell Extract, and in Cells by Electron Paramagnetic Resonance Distance Measurements'. Together they form a unique fingerprint.

Cite this