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Structural and functional characterization of human Iba proteins

Schulze, Jörg O ; Quedenau, Claudia ; Roske, Yvette ; Adam, Thomas ; Schüler, Herwig LU orcid ; Behlke, Joachim ; Turnbull, Andrew P ; Sievert, Volker ; Scheich, Christoph and Mueller, Uwe , et al. (2008) In The FEBS Journal 275(18). p.40-4627
Abstract

Iba2 is a homolog of ionized calcium-binding adapter molecule 1 (Iba1), a 17-kDa protein that binds and cross-links filamentous actin (F-actin) and localizes to membrane ruffles and phagocytic cups. Here, we present the crystal structure of human Iba2 and its homodimerization properties, F-actin cross-linking activity, cellular localization and recruitment upon bacterial invasion in comparison with Iba1. The Iba2 structure comprises two central EF-hand motifs lacking bound Ca2+. Iba2 crystallized as a homodimer stabilized by a disulfide bridge and zinc ions. Analytical ultracentrifugation revealed a different mode of dimerization under reducing conditions that was independent of Ca2+. Furthermore, no binding of Ca2+ up to 0.1 mM was... (More)

Iba2 is a homolog of ionized calcium-binding adapter molecule 1 (Iba1), a 17-kDa protein that binds and cross-links filamentous actin (F-actin) and localizes to membrane ruffles and phagocytic cups. Here, we present the crystal structure of human Iba2 and its homodimerization properties, F-actin cross-linking activity, cellular localization and recruitment upon bacterial invasion in comparison with Iba1. The Iba2 structure comprises two central EF-hand motifs lacking bound Ca2+. Iba2 crystallized as a homodimer stabilized by a disulfide bridge and zinc ions. Analytical ultracentrifugation revealed a different mode of dimerization under reducing conditions that was independent of Ca2+. Furthermore, no binding of Ca2+ up to 0.1 mM was detected by equilibrium dialysis. Correspondingly, Iba EF-hand motifs lack residues essential for strong Ca2+ coordination. Sedimentation experiments and microscopy detected pronounced, indistinguishable F-actin binding and cross-linking activity of Iba1 and Iba2 with induction of F-actin bundles. Fluorescent Iba fusion proteins were expressed in HeLa cells and co-localized with F-actin. Iba1 was recruited into cellular projections to a larger extent than Iba2. Additionally, we studied Iba recruitment in a Shigella invasion model that induces cytoskeletal rearrangements. Both proteins were recruited into the bacterial invasion zone and Iba1 was again concentrated slightly higher in the cellular extensions.

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publishing date
type
Contribution to journal
publication status
published
keywords
Actins/metabolism, Amino Acid Sequence, Calcium/metabolism, Calcium-Binding Proteins/analysis, Crystallography, X-Ray, DNA-Binding Proteins/analysis, Dimerization, HeLa Cells, Humans, Microfilament Proteins/analysis, Models, Molecular, Molecular Sequence Data, Sequence Alignment, Shigella/pathogenicity
in
The FEBS Journal
volume
275
issue
18
pages
14 pages
publisher
John Wiley & Sons Inc.
external identifiers
  • scopus:50849112730
  • pmid:18699778
ISSN
1742-464X
DOI
10.1111/j.1742-4658.2008.06605.x
language
English
LU publication?
no
id
f640a50e-2c0d-4653-bca0-52a22055a859
date added to LUP
2024-11-21 18:03:54
date last changed
2025-07-05 12:34:36
@article{f640a50e-2c0d-4653-bca0-52a22055a859,
  abstract     = {{<p>Iba2 is a homolog of ionized calcium-binding adapter molecule 1 (Iba1), a 17-kDa protein that binds and cross-links filamentous actin (F-actin) and localizes to membrane ruffles and phagocytic cups. Here, we present the crystal structure of human Iba2 and its homodimerization properties, F-actin cross-linking activity, cellular localization and recruitment upon bacterial invasion in comparison with Iba1. The Iba2 structure comprises two central EF-hand motifs lacking bound Ca2+. Iba2 crystallized as a homodimer stabilized by a disulfide bridge and zinc ions. Analytical ultracentrifugation revealed a different mode of dimerization under reducing conditions that was independent of Ca2+. Furthermore, no binding of Ca2+ up to 0.1 mM was detected by equilibrium dialysis. Correspondingly, Iba EF-hand motifs lack residues essential for strong Ca2+ coordination. Sedimentation experiments and microscopy detected pronounced, indistinguishable F-actin binding and cross-linking activity of Iba1 and Iba2 with induction of F-actin bundles. Fluorescent Iba fusion proteins were expressed in HeLa cells and co-localized with F-actin. Iba1 was recruited into cellular projections to a larger extent than Iba2. Additionally, we studied Iba recruitment in a Shigella invasion model that induces cytoskeletal rearrangements. Both proteins were recruited into the bacterial invasion zone and Iba1 was again concentrated slightly higher in the cellular extensions.</p>}},
  author       = {{Schulze, Jörg O and Quedenau, Claudia and Roske, Yvette and Adam, Thomas and Schüler, Herwig and Behlke, Joachim and Turnbull, Andrew P and Sievert, Volker and Scheich, Christoph and Mueller, Uwe and Heinemann, Udo and Büssow, Konrad}},
  issn         = {{1742-464X}},
  keywords     = {{Actins/metabolism; Amino Acid Sequence; Calcium/metabolism; Calcium-Binding Proteins/analysis; Crystallography, X-Ray; DNA-Binding Proteins/analysis; Dimerization; HeLa Cells; Humans; Microfilament Proteins/analysis; Models, Molecular; Molecular Sequence Data; Sequence Alignment; Shigella/pathogenicity}},
  language     = {{eng}},
  number       = {{18}},
  pages        = {{40--4627}},
  publisher    = {{John Wiley & Sons Inc.}},
  series       = {{The FEBS Journal}},
  title        = {{Structural and functional characterization of human Iba proteins}},
  url          = {{http://dx.doi.org/10.1111/j.1742-4658.2008.06605.x}},
  doi          = {{10.1111/j.1742-4658.2008.06605.x}},
  volume       = {{275}},
  year         = {{2008}},
}