Epitopes of adhesion-perturbing monoclonal antibodies map within a predicted alpha-helical domain of the integrin beta 1 subunit

1997 ◽  
Vol 110 (20) ◽  
pp. 2619-2628
Author(s):  
D.T. Shih ◽  
D. Boettiger ◽  
C.A. Buck
Keyword(s):  
Ligand Binding ◽  
Critical Role ◽  
Alpha Helix ◽  
Cation Binding ◽  
Affinity Column ◽  
Integrin Subunit ◽  
Amino Acid Residues ◽  
Helix Formation ◽  
Ligand Interactions ◽  
Integrin Ligand

Several recent studies have demonstrated the involvement of various domains of the beta 1 integrin subunit in ligand binding. Thus, specific amino acids have been shown to be important in divalent cation binding, and others have been implicated by peptide crosslinking to play an intimate role in integrin-ligand interactions. Added to these data are previous observations that a group of adhesion-blocking anti-chicken beta 1 antibodies mapped within the first 160 amino acid residues of the subunit. These observations suggested that this region plays a critical role in integrin ligand recognition. In order to further define the domain in which the epitopes for these antibodies are clustered, a series of mouse/chicken chimeric beta 1 constructs were examined for their reactivity with each of these antibodies. Most of the antibodies recognize a region between residues 124 to 160 of the chicken beta 1 subunit. Computer modeling predicted a possible amphipathic alpha-helical configuration for the region between residues 141 to 160. Consistent with this prediction, circular dichroism and NMR analysis revealed a tendency for a synthetic peptide containing these residues to form an alpha-helix. The significance of this structural characteristic was demonstrated by a mutation at residue 149 that disrupted the alpha-helix formation and resulted in a loss of the ability to form heterodimers with alpha subunits, localize to focal contacts, or be transported to the cell surface. The direct involvement of residues 141 to 160 in ligand binding was supported by the ability of a peptide with this sequence to elute integrins from a fibronectin affinity column. Thus, our data suggest that residues 141 to 160 of the integrin beta 1 subunit, when arranged in an alpha-helix configuration, participate in ligand binding.

scholarly journals Helix-coil transition theories. Are they correct?

1997 ◽  
Vol 44 (3) ◽  
pp. 423-432 ◽  
Author(s):  
A Bierzyński ◽  
K Pawłowski
Keyword(s):  
Water Solution ◽  
Alpha Helix ◽  
Theoretical Description ◽  
Random Coil ◽  
Specific Situation ◽  
Amino Acid Residues ◽  
Helix Formation ◽  
Helix Nucleation ◽  
Propagation Parameters ◽  
Helix Propagation

Principles of contemporary theoretical description of alpha-helix formation by polypeptide chains in water solution are shortly presented and critically discussed. The theory treats the unfolded state of a peptide as "random coil"--an ideal conformation quite distant from reality. We suggest that for this reason the helix propagation parameters of amino-acid residues, determined using series of model peptides with different sequential patterns, are not the same. Interpretation of the so called "nucleation parameter" is erroneous. In fact, it is not determined by the helix nucleation process but rather by a specific situation of residues at the helix N- and C-termini, and it strongly depends on solvation of their NH and CO groups, respectively. Consequently, helical segments with terminal sequences dominated by residues with strongly hydrophobic, bulky side chains can be very unstable. We postulate that an unexpectedly high stability of very short, pre-nucleated helices studied by us arises from a "helix end separation effect": separated helix termini are better solvated than when they overlap each other. Because of this effect, helix initiation may be much more difficult than predicted by the theoretical "helix nucleation parameters".

scholarly journals Integrin α4 cysteines 278 and 717 modulate VLA-4 ligand binding and also contribute to α4/180 formation

1996 ◽  
Vol 313 (3) ◽  
pp. 899-908 ◽  
Author(s):  
Cristina PUJADES ◽  
Joaquin TEIXIDÓ ◽  
Gianfranco BAZZONI ◽  
Martin E. HEMLER
Keyword(s):  
Cell Adhesion ◽  
Ligand Binding ◽  
Divalent Cations ◽  
Ligand Interactions ◽  
Integrin Α4 ◽  
Stringent Test ◽  
Integrin Ligand ◽  
Cell Adhesion Molecule 1 ◽  
Adhesion Ligand ◽  
Α Subunits

Here we describe experiments in which we mutated four of the six integrin α4 subunit cysteine residues that are not present in most other integrin α subunits that lack an I domain. In four different types of ligand binding assay we found that optimal integrin α4β1 (VLA-4) binding to vascular cell adhesion molecule 1 (VCAM-1) and/or to CS1 peptide required the presence of both α4 Cys278 and Cys717. In addition, optimal ligand binding required divalent cations and reduced cysteines, as evidenced by EDTA and N-ethylmaleimide inhibition results. In a control experiment, an α4 mutation that completely eliminated the α4 80/70 proteolytic cleavage site had no effect on ligand binding. Notably, although Cys278 and Cys717 mutations markedly altered ligand binding, they had no adverse effect on cell adhesion. Thus, compared with cell adhesion, ligand binding is a distinct and apparently more stringent test of VLA-4 integrin–ligand interactions. In addition, we have established that the formation of the previously described α4/180 [Parker, Pujades, Brenner and Hemler (1993) J. Biol. Chem. 268, 7028–7035] also requires Cys278 and Cys717, divalent cations and reduced cysteines. Thus α4/180 appears to be more functionally relevant than α4/150.

scholarly journals The structure of SALSA/DMBT1 SRCR domains reveal the conserved ligand-binding mechanism of the ancient SRCR-fold

2019 ◽  
Author(s):  
Martin P. Reichhardt ◽  
Vuokko Loimaranta ◽  
Susan M. Lea ◽  
Steven Johnson
Keyword(s):  
Ligand Binding ◽  
Scavenger Receptor ◽  
Cation Binding ◽  
Binding Mechanism ◽  
Ligand Interaction ◽  
Srcr Domain ◽  
Binding Interface ◽  
Cation Binding Site ◽  
Ligand Interactions ◽  
High Level

AbstractThe scavenger receptor cysteine-rich (SRCR) family of proteins comprise more than 20 membrane-associated and secreted molecules. Characterised by the presence of one or more copies of the ~110 amino acid SRCR domain, this class of proteins have widespread functions as anti-microbial molecules, scavenger- and signalling-receptors. Despite the high level of structural conservation of SRCR domains, no molecular basis for ligand interaction has been described. The SRCR protein SALSA, also known as dmbt1/gp340, is a key player in mucosal immunology. Based on detailed structures of the SALSA SRCR domains 1 and 8, we here reveal a novel universal ligand binding mechanism for SALSA ligands. The binding interface incorporates a dual cation binding site, which is highly conserved across the SRCR super family. Along with the well-described cation dependency on most SRCR domain-ligand interactions, our data suggest that the binding mechanism described for the SALSA SRCR domains is applicable to all SRCR domains. We thus propose to have identified in SALSA a conserved functional mechanism for ligand recognition by the SRCR class of proteins.

scholarly journals Cyclic Mechanical Reinforcement of Integrin–Ligand Interactions

2013 ◽  
Vol 49 (6) ◽  
pp. 1176 ◽  
Author(s):  
Fang Kong ◽  
Zhenhai Li ◽  
William M. Parks ◽  
David W. Dumbauld ◽  
Andrés J. García ◽  
...  
Keyword(s):  
Mechanical Reinforcement ◽  
Ligand Interactions ◽  
Integrin Ligand

ALPHA-HELIX FORMATION IN C-PEPTIDE RNASE-A INVESTIGATED BY PARALLEL TEMPERING SIMULATIONS

2007 ◽  
Vol 18 (01) ◽  
pp. 91-98 ◽  
Author(s):  
GÖKHAN GÖKOĞLU ◽  
TARIK ÇELİK
Keyword(s):  
Energy State ◽  
Minimum Energy ◽  
Alpha Helix ◽  
Salt Bridge ◽  
Rnase A ◽  
Parallel Tempering ◽  
Implicit Solvent ◽  
C Peptide ◽  
Helix Formation ◽  
Α Helix

We have performed parallel tempering simulations of a 13-residue peptide fragment of ribonuclease-A, c-peptide, in implicit solvent with constant dielectric permittivity. This peptide has a strong tendency to form α-helical conformations in solvent as suggested by circular dichroism (CD) and nuclear magnetic resonance (NMR) experiments. Our results demonstrate that 5th and 8–12 residues are in the α-helical region of the Ramachandran map for global minimum energy state in solvent environment. Effects of salt bridge formation on stability of α-helix structure are discussed.

Conformational dynamics and thermodynamics of protein–ligand binding studied by NMR relaxation

2012 ◽  
Vol 40 (2) ◽  
pp. 419-423 ◽  
Author(s):  
Mikael Akke
Keyword(s):  
Ligand Binding ◽  
Bound States ◽  
Conformational Dynamics ◽  
Nmr Relaxation ◽  
Titration Calorimetry ◽  
Conformational Entropy ◽  
Chain Order ◽  
Ligand Interactions ◽  
Galectin 3 ◽  
Relaxation Experiments

Protein conformational dynamics can be critical for ligand binding in two ways that relate to kinetics and thermodynamics respectively. First, conformational transitions between different substates can control access to the binding site (kinetics). Secondly, differences between free and ligand-bound states in their conformational fluctuations contribute to the entropy of ligand binding (thermodynamics). In the present paper, I focus on the second topic, summarizing our recent results on the role of conformational entropy in ligand binding to Gal3C (the carbohydrate-recognition domain of galectin-3). NMR relaxation experiments provide a unique probe of conformational entropy by characterizing bond-vector fluctuations at atomic resolution. By monitoring differences between the free and ligand-bound states in their backbone and side chain order parameters, we have estimated the contributions from conformational entropy to the free energy of binding. Overall, the conformational entropy of Gal3C increases upon ligand binding, thereby contributing favourably to the binding affinity. Comparisons with the results from isothermal titration calorimetry indicate that the conformational entropy is comparable in magnitude to the enthalpy of binding. Furthermore, there are significant differences in the dynamic response to binding of different ligands, despite the fact that the protein structure is virtually identical in the different protein–ligand complexes. Thus both affinity and specificity of ligand binding to Gal3C appear to depend in part on subtle differences in the conformational fluctuations that reflect the complex interplay between structure, dynamics and ligand interactions.

SATP (YaaH), a succinate–acetate transporter protein in Escherichia coli

2013 ◽  
Vol 454 (3) ◽  
pp. 585-595 ◽  
Author(s):  
Joana Sá-Pessoa ◽  
Sandra Paiva ◽  
David Ribas ◽  
Inês Jesus Silva ◽  
Sandra Cristina Viegas ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Acetic Acid ◽  
Succinic Acid ◽  
Critical Role ◽  
Amino Acid Residues ◽  
E Coli ◽  
Transporter Protein ◽  
Affinity Constants ◽  
In Trans

In the present paper we describe a new carboxylic acid transporter in Escherichia coli encoded by the gene yaaH. In contrast to what had been described for other YaaH family members, the E. coli transporter is highly specific for acetic acid (a monocarboxylate) and for succinic acid (a dicarboxylate), with affinity constants at pH 6.0 of 1.24±0.13 mM for acetic acid and 1.18±0.10 mM for succinic acid. In glucose-grown cells the ΔyaaH mutant is compromised for the uptake of both labelled acetic and succinic acids. YaaH, together with ActP, described previously as an acetate transporter, affect the use of acetic acid as sole carbon and energy source. Both genes have to be deleted simultaneously to abolish acetate transport. The uptake of acetate and succinate was restored when yaaH was expressed in trans in ΔyaaH ΔactP cells. We also demonstrate the critical role of YaaH amino acid residues Leu131 and Ala164 on the enhanced ability to transport lactate. Owing to its functional role in acetate and succinate uptake we propose its assignment as SatP: the Succinate–Acetate Transporter Protein.

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