Effects of Metal-Binding Loop Mutations on Ligand Binding to Calcium- and Integrin-Binding Protein 1. Evolution of the EF-Hand?

Yamniuk, Aaron P.; Gifford, Jessica L.; Linse, Sara; Vogel, Hans J.

doi:10.1021/bi701494m

Cited by 13 publications

(17 citation statements)

References 48 publications

(86 reference statements)

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“…Although the C-terminal end (residues 158 -191) of Mg 2ϩ -CIB1 could only be partially assigned (18), the corresponding part of Mg 2ϩ -CIB1, once complexed with the ␣IIb peptide, can be almost fully assigned. Apparently, the presence of the ␣IIb peptide increases the binding affinity of the EF-IV loop for Mg 2ϩ resulting in the stabilization of the structure in the EF-IV region (29). Comparison of the ⌬R 2,eff of Mg 2ϩ -CIB1 and the ␣IIb-bound form of the protein reveals that H7 and H9 could be directly involved in the interaction with the ␣IIb peptide, which is consistent with the CSP results (Fig.…”

Section: Methyl Groups Probe the Interaction Between Cib1 And ␣Iib-supporting

confidence: 88%

“…This methyl assignment approach requires preliminary knowledge of the chemical shifts of C␣ and C␤, which have been previously obtained from the backbone assignment work (12,18,29 The backbone amide 15 N relaxation dispersion measurements were carried out using a CPMG relaxation dispersion experiment (21) implemented with an inter-scan delay of 2 s and a total constant CPMG length (T CP ) of 61.08 ms. Two sets of CT-CPMG experiments were acquired at CPMG field strengths ( CPMG ) of 50 and 500 Hz, respectively. For each CPMG field strength, two replicate data sets were collected each with 32 dummy scans and 64 scans per t1 point.…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Solution Structures of Ca2+-CIB1 and Mg2+-CIB1 and Their Interactions with the Platelet Integrin αIIb Cytoplasmic Domain

Huang

Ishida

Yamniuk

et al. 2011

Journal of Biological Chemistry

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The calcium-and integrin-binding protein 1 (CIB1) is a ubiquitous Ca 2؉ -binding protein and a specific binding partner for the platelet integrin ␣IIb cytoplasmic domain, which confers the key role of CIB1 in hemostasis. CIB1 is also known to be involved in apoptosis, embryogenesis, and the DNA damage response. In this study, the solution structures of both Ca 2؉ -CIB1 and Mg 2؉ -CIB1 were determined using solution-state NMR spectroscopy. The methyl groups of Ile, Leu, and Val were selectively protonated to compensate for the loss of protons due to deuteration. The solution structure of Ca 2؉ -CIB1 possesses smaller opened EF-hands in its C-domain compared with available crystal structures. Ca 2؉ -CIB1 and Mg 2؉ -CIB1 have similar structures, but the N-lobe of Mg 2؉ -CIB1 is slightly more opened than that of Ca 2؉ -CIB1. Additional NMR experiments, such as chemical shift perturbation and methyl group solvent accessibility as measured by a nitroxide surface probe, were carried out to further characterize the structures of Ca 2؉ -CIB1 and Mg 2؉ -CIB1 as well as their interactions with the integrin ␣IIb cytoplasmic domain. NMR measurements of backbone amide proton slow motion (microsecond to millisecond) dynamics confirmed that the C-terminal helix of Ca 2؉ -CIB1 is displaced upon ␣IIb binding. The EF-hand III of both Ca 2؉ -CIB1 and Mg 2؉ -CIB1 was identified to be directly involved in the interaction of CIB1 with ␣IIb. Together, these data illustrate that CIB1 behaves quite differently from related EF-hand regulatory calcium-binding proteins, such as calmodulin or neuronal calcium sensor proteins.The calcium-and integrin-binding protein 1 (CIB1) is a member of the regulatory Ca 2ϩ -binding helix-loop-helix or EFhand protein family (1). CIB1 is a ubiquitous 191-residue (22 kDa) protein with several functions in cell signaling (2, 3). CIB1 has been reported to interact with a number of protein targets, including the platelet integrin ␣IIb subunit (4), sphingosine kinase 1 (5), p21-activated kinase (6), apoptosis signal-regulating kinase (7), polo-like protein kinases (8), and a recently reported new target in the regulation of cardiac hypertrophy, calcineurin B (9). The interaction of CIB1 with the integrin ␣IIb subunit has been studied extensively (10 -13). The integrin ␣IIb subunit usually associates with the integrin ␤3 subunit, and the heterodimeric ␣IIb␤3 protein is involved in both so-called "inside-out" and "outside-in" signaling pathways (3). CIB1 is believed to be capable of specifically binding to the ␣IIb cytoplasmic domain and dissociating the ␣IIb␤3 dimer, which in turn triggers integrin activation (11,12). The interactions between Ca 2ϩ -CIB1 and a fragment of the integrin ␣IIb subunit (residues 983-1008, hereafter referred to as the ␣IIb peptide) have been suggested to mainly involve a hydrophobic pocket in the C-domain of Ca 2ϩ -CIB1 with a dissociation constant in the submicromolar range (10).The calcium-bound CIB1 (Ca 2ϩ -CIB1) structure consists of four helix-loop-helix (EF-hand) Ca 2ϩ -binding m...

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Section: Methyl Groups Probe the Interaction Between Cib1 And ␣Iib-supporting

confidence: 88%

Section: Methodsmentioning

confidence: 99%

Solution Structures of Ca2+-CIB1 and Mg2+-CIB1 and Their Interactions with the Platelet Integrin αIIb Cytoplasmic Domain

Huang

Ishida

Yamniuk

et al. 2011

Journal of Biological Chemistry

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“…Therefore, 1 H, 13 C-HSQC spectra for the methyl groups of CIB1 complexed with αIIb-L were recorded in the presence of either Ca 2+ or Mg 2+ (Figure S4B). Similar to the backbone NMR studies, 55 methyl side chain 1 H, 13 C-HSQC spectra for Ca 2+ - and Mg 2+ -bound CIB1/αIIb-L show nearly the same pattern with minor deviations except for two residues in the calcium/magnesium binding loop, I168 and L123 (Figure S4B). These results suggest that Ca 2+ -CIB and Mg 2+ -CIB1 interact with αIIb in nearly the same manner.…”

Section: Resultssupporting

confidence: 72%

Structural Basis for the Activation of Platelet Integrin αIIbβ3 by Calcium- and Integrin-Binding Protein 1

Huang

Vogel

2012

J. Am. Chem. Soc.

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Calcium and integrin binding protein 1 (CIB1) is a specific binding partner for the cytoplasmic domain of the αIIb subunit of the highly abundant platelet integrin αIIbβ3. This protein has been suggested to be involved in the regulation of the activation of αIIbβ3, a process leading to platelet aggregation and blood coagulation. In this work, the solution structure of the deuterated Ca2+-CIB1 protein complexed with an αIIb peptide was first determined through modern RDC-based NMR methods. Next, we generated a complex structure for CIB1 and the αIIb domain (Ca2+-CIB1/αIIb) using the program Haddock, which is based on experimental restraints obtained for the protein–peptide interface from cross-saturation NMR experiments. In this data-driven complex structure, the N-terminal α-helix of the cytoplasmic domain of αIIb is buried in the hydrophobic pocket of the C-lobe of Ca2+-CIB1. The C-terminal acidic tail of αIIb remains unstructured and likely interacts with several positively charged residues in the N-lobe of Ca2+-CIB1. A potential molecular mechanism for the CIB1-mediated activation of the platelet integrin could be proposed on the basis of the model structure of this protein complex. Another feature of this work is that, in the NMR cross-saturation experiments, we applied the selective radio frequency irradiation to the smaller binding partner (the αIIb peptide), and successfully detected the binding interface on the larger binding partner Ca2+-CIB1 through its selectively protonated methyl groups. This ‘reverse’ methodology has a broad potential to be employed to many other complexes where synthetic peptides and a suitably isotope-labeled medium- to large-sized protein are used to study protein–protein interactions.

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“…To gain insights into CIB1 function, the structure of CIB1 has been explored extensively by NMR, circular dichroism, X-ray crystallography, and sequence analyses (2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12). CIB1 is a small, 22-kDa, 191-aa, intracellular Ca 2+ -binding protein that possesses an N-terminal myristoylation site (3,13).…”

Section: Cib1 Structure and Functionmentioning

confidence: 99%

CIB1: a small protein with big ambitions

et al. 2016

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Calcium- and integrin-binding protein 1 (CIB1) is a small, ubiquitously expressed protein that was first identified as an intracellular binding partner of a platelet-specific α-integrin cytoplasmic tail. Although early studies revealed a role for CIB1 in regulating platelet integrin activity, recent studies have indicated a more diverse role for CIB1 in many different cell types and processes, including calcium signaling, migration, adhesion, proliferation, and survival. Increasing evidence also points to a novel role for CIB1 in cancer and cardiovascular disease. In addition, an array of CIB1 binding partners has been identified that provide important insight into how CIB1 may regulate these processes. Some of these binding partners include the serine/threonine kinases, p21-activated kinase 1 (PAK1), apoptosis signal-regulating kinase 1 (ASK1), and polo-like kinase 3 (PLK3). Structural and mutational studies indicate that CIB1 binds most or all of its partners via a well-defined hydrophobic cleft. Although CIB1 itself lacks known enzymatic activity, it supports the PI3K/AKT and MEK/ERK oncogenic signaling pathways, in part, by directly modulating enzymes in these pathways. In this review, we discuss our current understanding of CIB1 and key questions regarding structure and function and how this seemingly diminutive protein impacts important signaling pathways and cellular processes in human health and disease.-Leisner, T. M., Freeman, T. C., Black, J. L., Parise, L. V. CIB1: a small protein with big ambitions.

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Effects of Metal-Binding Loop Mutations on Ligand Binding to Calcium- and Integrin-Binding Protein 1. Evolution of the EF-Hand?

Cited by 13 publications

References 48 publications

Solution Structures of Ca2+-CIB1 and Mg2+-CIB1 and Their Interactions with the Platelet Integrin αIIb Cytoplasmic Domain

Solution Structures of Ca2+-CIB1 and Mg2+-CIB1 and Their Interactions with the Platelet Integrin αIIb Cytoplasmic Domain

Structural Basis for the Activation of Platelet Integrin αIIbβ3 by Calcium- and Integrin-Binding Protein 1

CIB1: a small protein with big ambitions

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