Double Electron-Electron Resonance Probes Ca2+-Induced Conformational Changes and Dimerization of Recoverin
(2013) In Biochemistry 52(34). p.5800-5808- Abstract
- Recoverin a member of the neuronal calcium sensor (NCS) branch of the calmodulin superfamily, is expressed in retinal photoreceptor cells and serves as a calcium sensor in vision. Ca2+-induced conformational changes in recoverin cause extrusion of its covalently attached myristate (termed Ca2+-myristoyl switch) that promotes translocation of recoverin to disk membranes during phototransduction in retinal rod cells. Here we report double electron electron resonance (DEER) experiments on recoverin that probe Ca2+-induced changes in distance as measured by the dipolar coupling between spin-labels strategically positioned at engineered cysteine residues on the protein surface. The DEER distance between nitroxide spin-labels attached at C39 and... (More)
- Recoverin a member of the neuronal calcium sensor (NCS) branch of the calmodulin superfamily, is expressed in retinal photoreceptor cells and serves as a calcium sensor in vision. Ca2+-induced conformational changes in recoverin cause extrusion of its covalently attached myristate (termed Ca2+-myristoyl switch) that promotes translocation of recoverin to disk membranes during phototransduction in retinal rod cells. Here we report double electron electron resonance (DEER) experiments on recoverin that probe Ca2+-induced changes in distance as measured by the dipolar coupling between spin-labels strategically positioned at engineered cysteine residues on the protein surface. The DEER distance between nitroxide spin-labels attached at C39 and N120C is 2.5 +/- 0.1 nm for Ca2+-free recoverin and 3.7 +/- 0.1 nm for Ca2+-bound recoverin. An additional DEER distance (5-6 nm) observed for Ca2+-bound recoverin may represent an intermolecular distance between C39 and N120. N-15 NMR relaxation analysis and CW-EPR experiments both confirm that Ca2+-bound recoverin forms a dimer at protein concentrations above 100 mu M, whereas Ca2+-free recoverin is monomeric We propose that Ca2+-induced dimerization of recoverin at the disk membrane surface may play a role in regulating Ca2+-dependent phosphorylation of dimeric rhodopsin. The DEER approach will be useful for elucidating dimeric structures of NCS proteins in general for which Ca2+-induced dimerization is functionally important but not well understood. (Less)
Please use this url to cite or link to this publication:
https://lup.lub.lu.se/record/4062932
- author
- Myers, William K. ; Xu, Xianzhong ; Li, Congmin ; Lagerstedt, Jens LU ; Budamagunta, Madhu S. ; Voss, John C. ; Britt, R. David and Ames, James B.
- organization
- publishing date
- 2013
- type
- Contribution to journal
- publication status
- published
- subject
- in
- Biochemistry
- volume
- 52
- issue
- 34
- pages
- 5800 - 5808
- publisher
- The American Chemical Society (ACS)
- external identifiers
-
- wos:000323810900010
- scopus:84883238526
- pmid:23906368
- ISSN
- 0006-2960
- DOI
- 10.1021/bi400538w
- language
- English
- LU publication?
- yes
- id
- 4eabec65-41e0-4828-aa8b-c93e7ecaf245 (old id 4062932)
- date added to LUP
- 2016-04-01 10:11:13
- date last changed
- 2022-01-25 20:37:05
@article{4eabec65-41e0-4828-aa8b-c93e7ecaf245,
abstract = {{Recoverin a member of the neuronal calcium sensor (NCS) branch of the calmodulin superfamily, is expressed in retinal photoreceptor cells and serves as a calcium sensor in vision. Ca2+-induced conformational changes in recoverin cause extrusion of its covalently attached myristate (termed Ca2+-myristoyl switch) that promotes translocation of recoverin to disk membranes during phototransduction in retinal rod cells. Here we report double electron electron resonance (DEER) experiments on recoverin that probe Ca2+-induced changes in distance as measured by the dipolar coupling between spin-labels strategically positioned at engineered cysteine residues on the protein surface. The DEER distance between nitroxide spin-labels attached at C39 and N120C is 2.5 +/- 0.1 nm for Ca2+-free recoverin and 3.7 +/- 0.1 nm for Ca2+-bound recoverin. An additional DEER distance (5-6 nm) observed for Ca2+-bound recoverin may represent an intermolecular distance between C39 and N120. N-15 NMR relaxation analysis and CW-EPR experiments both confirm that Ca2+-bound recoverin forms a dimer at protein concentrations above 100 mu M, whereas Ca2+-free recoverin is monomeric We propose that Ca2+-induced dimerization of recoverin at the disk membrane surface may play a role in regulating Ca2+-dependent phosphorylation of dimeric rhodopsin. The DEER approach will be useful for elucidating dimeric structures of NCS proteins in general for which Ca2+-induced dimerization is functionally important but not well understood.}},
author = {{Myers, William K. and Xu, Xianzhong and Li, Congmin and Lagerstedt, Jens and Budamagunta, Madhu S. and Voss, John C. and Britt, R. David and Ames, James B.}},
issn = {{0006-2960}},
language = {{eng}},
number = {{34}},
pages = {{5800--5808}},
publisher = {{The American Chemical Society (ACS)}},
series = {{Biochemistry}},
title = {{Double Electron-Electron Resonance Probes Ca2+-Induced Conformational Changes and Dimerization of Recoverin}},
url = {{http://dx.doi.org/10.1021/bi400538w}},
doi = {{10.1021/bi400538w}},
volume = {{52}},
year = {{2013}},
}