T cell receptor-ligand interactions: A conformational preequilibrium or an induced fit

Gakamsky, Dmitry M.; Luescher, Immanuel F.; Pecht, Israel

doi:10.1073/pnas.0402840101

Cited by 36 publications

(36 citation statements)

References 31 publications

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“…Results of the current SPR measurements and of our earlier studies (18) show that, as in all other investigations of TCRpMHC interactions by this method, the TCR CMV -pp65-HLA-A*0201 association rate constants are slower than those expected for diffusion-controlled reactions in solution, i.e., these do not reflect the actual elementary steps of the process. This finding is not surprising in view of the limited time resolution of the method.…”

Section: Discussionmentioning

confidence: 71%

“…1). The rate constants were calculated by using a two-species model accounting for known slight analyte heterogeneity (18). The association and dissociation rate constants of the major fraction (k on 1 and k off 2 ) as well as the equilibrium dissociation constants (K d ) are listed in Table 1.…”

Section: Tcr Specific For Cytomegalovirus Peptide (Tcrcmv)-pp65-hla-amentioning

confidence: 99%

“…However, practically all reports suggesting the operation of a two-step mechanism lack experimental evidence because the elementary steps of the TCR-ligand interactions were not resolved. This lack of information has therefore been, and remains, a significant experimental challenge (18). Here we report resolution of a two-phase interaction time course of the TCR and pMHC association monitored by stopped-flow measurements using FRET between a donor labeled specific ligand and its acceptor labeled TCR.…”

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confidence: 99%

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Kinetic evidence for a ligand-binding-induced conformational transition in the T cell receptor

Gakamsky

Lewitzki

Grell

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

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Thermodynamics and kinetics of the interaction between T cell receptor specific for cytomegalovirus peptide (TCRCMV) and its specific ligand, pp65-HLA-A*0201 complex, were studied by surface plasmon resonance and stopped-flow methods. In the latter measurements, fluorescence resonance energy transfer (FRET) between fluorescently labeled reactants was used. Thermodynamic data derived from surface plasmon resonance measurements suggest that the complex formation is driven by both favorable enthalpy and entropy. Two reaction phases were resolved by the stopped-flow measurements. The rate constant of the first step was calculated to be close to the diffusion-controlled limit rate (3⅐10 5 to 10 6 M ؊1 ⅐s ؊1 ), whereas the second step's reaction rate was found to be concentration independent and relatively slow (2-4 s ؊1 at 25°C). These findings strongly suggest that the interactions between the TCR and its ligand, the peptide-MHC complex, proceed by a two-step mechanism, in which the second step is an induced-fit process, rate determining for antigen recognition by TCR.conformational changes ͉ T cell receptor-ligand interactions ͉ FRET ͉ stopped flow ͉ induced fit T cell receptor (TCR) association with its ligand, a peptide bound to molecules encoded by class I or II of the major histocompatibility complexes (pMHC), is the initial process in T cell activation (1-3). This process can induce different types of immune responses depending on the nature of the pMHC and the involved T cell (4-6). It is well established that despite the prevalent micromolar affinity, this interaction is characterized by such an exquisite sensitivity that even just a few antigenic pMHC molecules can be recognized on an antigen-presenting cell (APC) (7). At the same time, a TCR was shown to cross-react with different antigenic pMHC ligands (8 -10). Threedimensional structures determined for more than two dozen different TCR-pMHC complexes revealed similar diagonal orientations of TCR molecules with respect to their pMHC ligands (11,12). It was also found that two of the complementaritydetermining regions (CDRs) of the TCR, CDR1 and CDR2, interact predominantly with the MHC molecules, whereas the CDR3 interacts with exposed peptide residues. Importantly, significant conformational changes in the TCR, mainly in its CDR3, have been observed in crystallographic studies (10,13,14). These structural studies provide a static picture and require time-resolved analysis of the process to deepen the understanding of the molecular mechanism of antigen recognition by the TCR.Kinetic studies carried out to date have used the surface plasmon resonance (SPR) method, which has several deficiencies. First and foremost, its time resolution is limited. Essentially, results of all such studies were interpreted as bimolecular processes with a relatively slow rate constant of complex formation and a fast rate of its dissociation (3). This slow association rate was assigned to a conformational transition in the TCR binding site (15). This notion provided the basis f...

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Section: Discussionmentioning

confidence: 71%

Section: Tcr Specific For Cytomegalovirus Peptide (Tcrcmv)-pp65-hla-amentioning

confidence: 99%

mentioning

confidence: 99%

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Kinetic evidence for a ligand-binding-induced conformational transition in the T cell receptor

Gakamsky

Lewitzki

Grell

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

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“…A rather good agreement between TCR-pMHC kinetic data and these two models has been established [19]. Thus, the induced-fit model suggests that the association reaction starts by forming an initial intermediate complex (TCRMHCp) int at a rate close to a diffusion-controlled process.…”

Section: Tcr-pmhc Associationmentioning

confidence: 73%

Spatial coordination of CD8 and TCR molecules controls antigen recognition by CD8⁺ T‐cells

Pecht

Gakamsky

2005

FEBS Letters

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The interactions between the TCR and peptides bound to class I MHC encoded molecules (pMHC) and a mechanism for CD8 cooperation in this process are reviewed. Observation of two TCR/CD8 populations with different lateral diffusion rate constants as well as two distinct association phases of class I MHC tetramers ((pMHC) 4 ) with T-cells suggest that the most efficient pMHC-T-cell association route corresponds to a fast tetramer binding to a colocalized CD8/TCR population, which apparently resides within membrane rafts. Thus, ligandcell association starts by pMHC binding to the CD8. This rather fast step promotes pMHC association with CD8-proximal TCRs and thereby enhances the overall association process. The model suggests that this raft-associated CD8-TCR subpopulation is responsible for evoking T-cell activation.

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“…As the global fitting routine of the evaluation software did not produce satisfactory results, the association and dissociation time courses were separately fitted to a single-exponential model, as described in Ref. 28, using Prism 4 (GraphPad Software, San Diego, CA). The influence of glucose and GKAs on the dissociation of the preformed GK-GKRP complex was tested using the regeneration scout tool provided by Biacore S51 control software.…”

Section: Methodsmentioning

confidence: 99%

Biophysical Characterization of the Interaction between Hepatic Glucokinase and Its Regulatory Protein

Anderka

Boyken

Aschenbach

et al. 2008

Journal of Biological Chemistry

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Glucokinase (GK) is a key enzyme of glucose metabolism in liver and pancreatic ␤-cells, and small molecule activators of GK (GKAs) are under evaluation for the treatment of type 2 diabetes. In liver, GK activity is controlled by the GK regulatory protein (GKRP), which forms an inhibitory complex with the enzyme. Here, we performed isothermal titration calorimetry and surface plasmon resonance experiments to characterize GK-GKRP binding and to study the influence that physiological and pharmacological effectors of GK have on the protein-protein interaction. In the presence of fructose-6-phosphate, GK-GKRP complex formation displayed a strong entropic driving force opposed by a large positive enthalpy; a negative change in heat capacity was observed (K d ‫؍‬ 45 nM, ⌬H ‫؍‬ 15.6 kcal/mol, T⌬S ‫؍‬ 25.7 kcal/mol, ⌬C p ‫؍‬ ؊354 cal mol ؊1 K ؊1 ). With k off ‫؍‬ 1.3 ؋ 10 ؊2 s ؊1 , the complex dissociated quickly. The thermodynamic profile suggested a largely hydrophobic interaction. In addition, effects of pH and buffer demonstrated the coupled uptake of one proton and indicated an ionic contribution to binding. Glucose decreased the binding affinity between GK and GKRP. This decrease was potentiated by an ATP analogue. Prototypical GKAs of the amino-heteroaryl-amide type bound to GK in a glucose-dependent manner and impaired the association of GK with GKRP. This mechanism might contribute to the antidiabetic effects of GKAs. Glucokinase (GK)2 is the predominant glucose-phosphorylating enzyme in liver and pancreatic ␤-cells and plays a central role in blood glucose homeostasis (1, 2). Enhancing GK activity by small molecule GK activators (GKAs) is currently under evaluation as an approach for the treatment of diabetes (3). Hepatic GK activity is controlled by an endogenous inhibitor, a 68-kDa GK regulatory protein (GKRP) (4). During starvation, the enzyme is bound to GKRP, leading to its inactivation and sequestration in the nucleus. After refeeding, the GK-GKRP complex dissociates and GK translocates into the cytoplasm (5, 6). In a rodent model of type 2 diabetes, this translocation is impaired, which could contribute to the defective blood glucose homeostasis (7). Further evidence for the metabolic impact of GKRP comes from recent human genome-wide association studies that show a strong linkage between a GKRP gene polymorphism and serum triglyceride levels (8, 9). The formation of the GK-GKRP complex is favored by fructose-6-phosphate (F6P) and inhibited by fructose-1-phosphate (F1P) (10). Both sugar phosphates bind to the same site on the regulatory protein (11). An increase of the intracellular F1P concentration leading to the release of GK from its inhibitory complex with GKRP is most likely the reason for the stimulation of hepatic glucose metabolism by catalytic amounts of fructose or sorbitol (6, 12, 13). Site-directed mutagenesis experiments indicate that the binding interface for GKRP lies close to the binding site of allosteric GKAs of the amino-heteroaryl-amide type (14). However, it is controversial if thes...

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T cell receptor-ligand interactions: A conformational preequilibrium or an induced fit

Cited by 36 publications

References 31 publications

Kinetic evidence for a ligand-binding-induced conformational transition in the T cell receptor

Kinetic evidence for a ligand-binding-induced conformational transition in the T cell receptor

Spatial coordination of CD8 and TCR molecules controls antigen recognition by CD8⁺ T‐cells

Biophysical Characterization of the Interaction between Hepatic Glucokinase and Its Regulatory Protein

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T cell receptor-ligand interactions: A conformational preequilibrium or an induced fit

Cited by 36 publications

References 31 publications

Kinetic evidence for a ligand-binding-induced conformational transition in the T cell receptor

Kinetic evidence for a ligand-binding-induced conformational transition in the T cell receptor

Spatial coordination of CD8 and TCR molecules controls antigen recognition by CD8+ T‐cells

Biophysical Characterization of the Interaction between Hepatic Glucokinase and Its Regulatory Protein

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Spatial coordination of CD8 and TCR molecules controls antigen recognition by CD8⁺ T‐cells