Faculty of 1000 evaluation for An internal ligand-bound, metastable state of a leukocyte integrin, αXβ2 (original) (raw)
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Structure and allosteric regulation of the X 2 integrin I domain
Proceedings of the National Academy of Sciences, 2003
The integrin ␣X2 (CD11c͞CD18, p150,95) binds ligands through the I domain of the ␣X subunit. Ligands include the complement factor fragment iC3b, a key component in the innate immune defense, which, together with the expression of ␣X2 on dendritic cells and on other leukocytes, suggests a role in the immune response. We now report the structure of the ␣X I domain resolved at 1.65 Å by x-ray crystallography. To analyze structural requirements for ligand binding we made a mutation in the ␣X I domain C-terminal helix, which increased the affinity for iC3b Ϸ200-fold to 2.4 M compared with the wild-type domain affinity of Ϸ400 M.
The structural bases of integrin-ligand interactions
Trends in Cell Biology, 1994
noncovalently associated u and [~ subunits. At least bases ul-ct and 8 13 subunits have been identified. On the basis of the subunits [3~, [32 and [33, the integrins were originally divided into three subfamilies: the [3~ and integrin ligand [33 subfamilies are inv°lved primarily in interacti°ns with ECM proteins, whereas the 132 subfamily, which . . is restricted to leukocytes, is also important for interactions cell-cell contact. The [31 subunit can combine ,,ith 10 et subunits (ot~ to ct 9, and cx~) to form heterodimers that specifically bind to ECM constituents such as fibronectin, laminin and different collagen types. The two major integrins of the [3 3 subfamily, U.b[33 and av[33, are mainly directed against vascular li-Many extracellular matrix (ECM) proteins, particularly those in gands, such as fibronectin, vitronectin, fibrinogen, von Will brand factor, thrombospondin and others the vascular system, use their classical integrin-recognition motif (Table 1).
Structure and allosteric regulation of the αXβ2 integrin I domain
Proceedings of the National Academy of Sciences of the United States of America, 2003
The integrin ␣X2 (CD11c͞CD18, p150,95) binds ligands through the I domain of the ␣X subunit. Ligands include the complement factor fragment iC3b, a key component in the innate immune defense, which, together with the expression of ␣X2 on dendritic cells and on other leukocytes, suggests a role in the immune response. We now report the structure of the ␣X I domain resolved at 1.65 Å by x-ray crystallography. To analyze structural requirements for ligand binding we made a mutation in the ␣X I domain C-terminal helix, which increased the affinity for iC3b Ϸ200-fold to 2.4 M compared with the wild-type domain affinity of Ϸ400 M. Gel permeation chromatography supported a conformational change between the wild-type and mutated domains. Conservation of allosteric regulation in the ␣X I domain points to the functional importance of this phenomenon.
F1000 - Post-publication peer review of the biomedical literature, 2016
Background: Integrins undergo large conformational changes when ligand-bound. Results: Using nanodisc technology and EM, we observed Mn 2ϩ-activated ␣ IIb  3 integrin both alone and fibrin-bound. Conclusion: MnCl 2-activated ␣ IIb  3 integrin alone has a compact conformation, becoming fully upright and open when fibrin-bound. Significance: The first structure of membrane-embedded integrins bound to physiological substrate reveals the importance of integrin extension in macromolecular ligand binding. Increased ligand binding to integrin ("activation") underpins many biological processes, such as leukocyte trafficking, cell migration, host-pathogen interaction, and hemostasis. Integrins exist in several conformations, ranging from compact and bent to extended and open. However, the exact conformation of membrane-embedded, full-length integrin bound to its physiological macromolecular ligand is still unclear. Integrin ␣ IIb  3 , the most abundant integrin in platelets, has been a prototype for integrin activation studies. Using negative stain electron microscopy and nanodisc-embedding to provide a membranelike environment, we visualized the conformation of full-length ␣ IIb  3 in both a Mn 2؉-activated, ligand-free state and a Mn 2؉activated, fibrin-bound state. Activated but ligand-free integrins exist mainly in the compact conformation, whereas fibrinbound ␣ IIb  3 predominantly exists in a fully extended, headpiece open conformation. Our results show that membrane-embedded, full-length integrin adopts an extended and open conformation when bound to its physiological macromolecular ligand.
Integrin αIIbβ3: ligand interactions are linked to binding‐site remodeling
2006
This studyt ested theh ypothesis that high-affinity bindingo fm acromolecularl igands to the a IIbb 3 integrin is tightlyc oupled to binding-site remodeling,a ni nduced-fit processt hats hiftsaconformational equilibriumf romaresting toward an open receptor.I nteractionsb e tween a IIb b 3a nd twom odel ligands-echistatin,a6-kDarecombinant proteinwithanRGD integrin-targeting sequence,and fibrinogen's g -module, a3 0-kDar ecombinant proteinw ith aK QAGDVi ntegrin bindings ite-were measured by sedimentationvelocity, fluorescence anisotropy,a nd as olid-phase bindinga ssay,a nd modeledbymolecular graphics.Studyingechistatinvariants(R24A,R24K, D26A,D26E, D27W,D27F),wefound that electrostatic contacts with chargedr esiduesa tt he a IIb/b 3i nterface,r athert hann onpolarc ontacts, perturbt he conformation of ther esting integrin.A spartate 26,w hich interacts with then earbyM IDAS cation,w as essentialfor binding, as D26A andD26Ewereinactive. In contrast,R24Kwas fullyand R24A partly active, indicating that thepositivelycharged arginine 24 contributesto, butisnot required for, integrin recognition. Moreover,w ed emonstrated that priming-i.e.,e ctodomainc onformationalc hanges ando ligomerization inducedbyincubationat35°Cwiththe ligand-mimetic peptidecHarGD-promotes complexformation with fibrinogen's g -module. We also observed that the g -module'sflexiblecarboxy terminus wasnot required for a IIb b 3i ntegrinb inding.O ur studiesd ifferentiate primingl igands,w hich bind to ther esting receptor and perturbits conformation,fromregulated ligands, wherebinding-siteremodelingmustfirst occur. Echistatin's bindingenergyissufficienttorearrange thesubunit interface, butregulated ligandslikefibrinogenmustrely on primingt oo vercomec onformational barriers.
Structure and allosteric regulation of the alpha X beta 2 integrin I domain
Proceedings of the National Academy of Sciences of the United States of America, 2003
The integrin alpha X beta 2 (CD11c/CD18, p150,95) binds ligands through the I domain of the alpha X subunit. Ligands include the complement factor fragment iC3b, a key component in the innate immune defense, which, together with the expression of alpha X beta 2 on dendritic cells and on other leukocytes, suggests a role in the immune response. We now report the structure of the alpha X I domain resolved at 1.65 A by x-ray crystallography. To analyze structural requirements for ligand binding we made a mutation in the alpha X I domain C-terminal helix, which increased the affinity for iC3b approximately 200-fold to 2.4 microM compared with the wild-type domain affinity of approximately 400 microM. Gel permeation chromatography supported a conformational change between the wild-type and mutated domains. Conservation of allosteric regulation in the alpha X I domain points to the functional importance of this phenomenon.
Critical Residues for Ligand Binding in an I Domain-like Structure of the Integrin β1 Subunit
Journal of Biological Chemistry, 1996
Several integrin ␣ subunits have an inserted sequence of about 200 residues (the I or A domain) that is critical for ligand interactions. The presence of an I domain-like structure within the integrin  subunit has been proposed based on the similarity of the hydropathy profiles and the homology of sequences between the ␣ and  subunits. This study was designed to determine whether the region of the 1 subunit that includes residues 101-335 has the characteristics of an I domain. We found novel critical residues for ligand binding (Ser-132, Asn-224, Asp-226, Glu-229, Asp-233, Asp-267, and Asp-295, in addition to the previously reported Asp-130) using sitedirected mutagenesis. The critical residues for ligand binding are located in several of loop structures of the region (or in a potential loop between an ␣ helix and a  strand), which have been predicted using multiple secondary structure prediction methods. The data suggest that the  subunit has multiple disrupted critical oxygenated residues for ligand binding similar to those found in the ␣ I domain.
Cation binding to the integrin CD11b I domain and activation model assessment
1998
The integrin family of cell-surface receptors mediate cell adhesion through interactions with the extracellular matrix or other cell-surface receptors. The α chain of some integrin heterodimers includes an inserted 'I domain' of about 200 amino acids which binds divalent metal ions and is essential for integrin function. Lee et al. proposed that the I domain of the integrin CD11b adopts a unique 'active' conformation when bound to its counter receptor. In addition, they proposed that the lack of adhesion in the presence of Ca 2+ ion reflected the stabilization of an 'inactive' I-domain conformation. We set out to independently determine the structure of the CD11b I domain and to evaluate the structural effects of divalent ion binding to this protein.