|
|
|
|
|
|
|
||
Articles |
Laboratory of Cell Biology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892
 |
Abstract |
|---|
 Top Abstract Materials and MethodsResults and DiscussionReferences |
|---|
Key Words: major histocompatibility complex • crystallography • xeno-reactivity • transplantation • T cell receptor
Abbreviations used: β2m, β2-microglobulin; CDR, complementarity determining region.
The TCR, expressed on the surface of CD8+ CTL, recognizes specific class I MHC/peptide complexes on APC. The result of the recognition is typically lysis of the APC. The mature class I molecule is a ternary complex containing a glycoprotein heavy chain, a noncovalently associated light chain β2-microglobulin (β2m),1 and a small antigenic peptide (typically 8–10 residues). Crystal structures of class I/peptide complexes reveal that the peptide-binding region of class I is a cleft formed by two
Recent crystallographic studies of TCR/class I complexes (A6/HLA-A2/Tax, B7/HLA-A2/Tax, 2C/H-2Kb/ dEV8, and N15/H-2Kb/VSV8) (6–9) reveal a similar, diagonal orientation of the TCR over the peptide-binding cleft of each class I/peptide complex. The complementarity determining region (CDR) loops from each TCR (with the exception of CDR2β in A6 and CDR1β in B7) make contacts with both the peptide and the
Although a TCR may often appear to be highly specific for a given cognate ligand, there are numerous examples in the literature of T cell cross-reactivity (10). Cross-recognized ligands can consist of different peptides presented by the same class I molecule, the same peptide presented by different class I allotypes, or distinct class I/peptide complexes. The ability of a single TCR to engage different but related ligands, either optimally or suboptimally, is proposed to play an important role in many immune processes (11–13).
Biochemical and molecular biological studies have been conducted to characterize the molecular basis for T cell cross-reactivity. The most popular model proposed from these studies is the molecular mimicry model, where different class I/peptide complexes form similar (spatially equivalent) antigenic surfaces in shape, charge, or both (14, 15). A recent paper (16) suggests a modified molecular mimicry model, in which a critical local charge mimicry formed by two residues contribute to the recognition of syngeneic and allogeneic class I/peptide complexes by the same TCR.
Cross-reactivity of TCR for different MHC allotypes within the same species is termed allo-reactivity, and cross-reactivity for MHC across species is termed xeno-reactivity. Xeno-reactivity requires the pairing of incompatible class I and CD8 molecules as well as the participation of incompatible costimulatory molecules for activation, and as such is more complicated than allo-reactivity. However, at the level of TCR and class I/peptide binding, xeno-reactivity can be viewed as an extreme case of T cell allo- or cross-reactivity. The study of a xeno-reactive system may reveal a more general mechanism which covers the whole scope of T cell cross-reactivity.
The CD8+ CTL clone AHIII12.2 was derived from a C57BL/6 (H-2b) mouse injected with the human lymphoblastoid cell line JY (17). It engages in peptide-specific xeno-recognition of the human class I molecule HLA-A*0201 (designated A2) complexed with the peptide ALWGFFPVL (designated p1049). AHIII12.2 also recognizes murine class I H-2Db (designated Db) complexed with the synthetic peptide FAPGFFPYL (designated p1058) and closely related analogues (18). The sequences of both the peptides and class I molecules are significantly different between these two complexes. Nevertheless, they are both recognized by a single TCR. Experiments done using singly substituted peptides indicate that the TCR cross-recognition of these ligands cannot be explained simply by a common motif defined at the level of the peptide (18).
Mutagenesis studies, although powerful, sometimes are difficult to interpret and are unable to provide a complete picture of the biological events involved. Therefore, we have engaged in the present study, and have determined the crystal structures of A2/p1049 and Db complexed with p1027 (FAPGVFPYM) to 2.5 Å and 2.8 Å resolution, respectively. p1027 is a disubstituted analogue of p1058 and is an equally good agonist towards AHIII12.2 (data not shown). We chose p1027 rather than p1058 for the crystallographic study to maximize the primary sequence differences between the two class I/peptide complexes so that we can examine the extreme scope of T cell cross-reactivity. Db/p1027 describes crystallographically how class I binds a peptide containing a noncanonical residue at an anchor position. More importantly, this study reveals, at a near atomic resolution, the molecular surfaces of two class I/peptide complexes recognized by the same TCR. A comparison of the potential TCR contacting surfaces shows that these two complexes do not possess obvious similarity. We propose that TCR binds the same areas of each class I molecule, utilizing both common and different contact residues in these areas. It requires that these contacts, although not identical between the two complexes, generate sufficient binding affinity between TCR and these complexes to allow for further T cell activation. This, rather than a simple common surface, accounts for the recognition of these two complexes by the same TCR.
Synthetic Peptides.
Crystallization.
Data Collection and Processing.
The diffraction data were processed using the programs Denzo and Scalepack (21). Data statistics are shown in Table I. 
helices in the 1 and 2 domains of the heavy chain (for review see reference 1). Each specific class I molecule binds a large set of peptides through the interaction between the peptide termini and conserved residues in MHC (2, 3). The peptides bound to a specific class I allotype share two or more relatively invariant residues (the anchor residues). The anchor residues bind in specific pockets generated by the polymorphic residues in the class I heavy chain (2, 4, 5). The peptide terminal residues and the anchor residues have a predictable orientation, whereas other residues in each peptide usually have unique main chain conformations and side chain orientations corresponding to their specific sequences (3). As yet, there is not a set of rules to predict the conformation of a peptide based on its sequence and the class I allotype to which it binds. 1 and 2 helices of class I molecule, although the individual residues that are in contact with the CDRs are different in each class I. There are significant numbers of interactions between conserved class I residues and TCR, leading to the proposition that the orientation of TCR observed in these crystal structures is a general feature of most TCR/class I complexes. The conformation of the class I molecule remains the same both before and after TCR binding (6–9), although the center of the Tax peptide adjusts its conformation upon TCR binding in the A6/HLA-A2/Tax and B7/HLA-A2/ Tax structures. Conversely, the peptide retains the same conformation in the 2C/H2-Kb/dEV8 structure (except for one side chain reorientation), but the CDR loops of the TCR (especially CDR1 and CDR3 in the subunit) show large shifts relative to their positions in the structure of the uncomplexed 2C TCR. A similar comparison cannot be made for A6 and B7 because the unliganded structures are not known.
Materials and Methods
Top
Abstract
Materials and Methods
Results and Discussion
References
Preparation of A2/p1049 and Db/p1027.
The cDNAs for Db and murine β2m were gifts from Dr. Stanley G. Nathenson (Albert Einstein College of Medicine, New York, NY) (19). A new construct of Db was created to make it consistent with the length of the A2 construct (20). The new construct was generated by PCR amplifying residues 1–275 and was cloned into plasmid vector PLM1 (gifts from Dr. G. Verdine at Harvard University, Cambridge, MA). The protein was expressed in BL21pLysS cells (Invitrogen Corp.). Due to incorrect primer design, the first Gly residue of the heavy chain was left out, and three extra residues (ArgTrpGlu) were added to the COOH terminus of the heavy chain. The constructs of A2 heavy chain and human β2m were identical to those described in Garboczi (20). Large quantities of A2, Db, human β2m, and murine β2m were produced as inclusion bodies and purified as described previously (20).
Peptides used for crystallization were synthesized by the Peptide Synthesis Facility at the University of North Carolina at Chapel Hill (Chapel Hill, NC) and the National Cancer Institute at NIH (Bethesda, MD). All peptides were purified by reversed-phase HPLC to >95% purity, and the identities were confirmed by laser-desorption mass spectroscopy.
A2/p1049 and Db/p1027 were folded in vitro as described in Garboczi (20). The folded complexes were concentrated using an Amicon ultrafiltration cell and purified by HPLC gel filtration chromatography (Biosep-SEC-S2000; Phenomenex). These complexes were further purified on a FPLC mono Q column (Pharmacia Biotech). Both complexes were crystallized using the hanging–drop vapor–diffusion method. The reservoir solution for A2/p1049 contained 12–16% PEG6000 and 6% dioxane in 25 mM MES buffer, pH 6.5. The hanging drop for A2/p1049 was a 1:1 mixture of the reservoir solution and 10 mg/ ml protein in 25 mM MES, pH 6.5. The reservoir solution for Db/p1027 contained 25% PEG8000, 150 mM NaCl, 0.8 M Gly, and 6% DMSO in 25 mM MES buffer, pH 6.5. The hanging drop was a 1:1 mixture of the reservoir solution and the protein solution which contained 10 mg/ml protein and 0.8 M Gly in 25 mM MES buffer, pH 6.5. Microseeding was necessary for both complexes to obtain crystals suitable for crystallographic studies.
Crystals of A2/p1049 were stored in 25 mM MES, pH 6.5, containing 20% PEG6000. Crystals of Db/p1027 were stored in 25 mM MES, pH 6.5, containing 30% PEG8000 and 0.8 M Gly. These crystals were transferred directly into their corresponding storage buffer plus 30% glycerol, and were immediately placed in the cryo stream (generated by the Oxford Cryo System, and the temperature was set at 100 K). The diffraction data were collected on Rigaku R-Axis IIC using Cu K radiation.
|
Refinement of A2/p1049.
Computational refinement was performed using the program X-PLOR version 3.851 (24) and Refmac in CCP4 (25). The structure was first refined in X-PLOR using 8–2.5 Å resolution data. Twofold noncrystallographic- symmetry restraints were applied to the two molecules in the asymmetric unit of HLA-A2/p1049 crystals with a weight of 300 kcal mol–1 Å–2. Overall isotropic B factor refinement was performed first, then rigid body refinement was carried out using three domains: 1/2, 3, and β2m. Overall anisotropic B factor refinement slightly improved Rfree values. Cycles of computational refinement using positional refinement and simulated annealing (26) followed by manual model rebuilding using maps with coefficients 2fo-fc and fo-fc (after density modification using DM) were carried out. The X-PLOR refined structure was further refined using Refmac with strict noncrystallographic-symmetry restraints (see Refmac manual), a bulk solvent correction, overall anisotropic B factor correction, and individual B factor refinement. 50 water molecules were added in the structure using program ARP (27) in CCP4. The final Rfree and Rwork factors (F > 0) between 30 and 2.5 Å resolution are 30.4 and 25.5%, respectively. The refinement statistics are shown in Table I. The final real-space correlation coefficient (23) between the refined model and the DM-modified electron density map with coefficients 2fo-fc is 0.76. The conformation of the peptide was confirmed by the following averaged omit map procedure (Fig. 1 A). Masks including peptide were generated, but the peptide was omitted from the refined model to generate the phases for the initial map. Twofold averaging, solvent flattening, and histogram matching were applied with the program DM to generate the final omit map.
|
All residues on the 1/2 helices have clear and unambiguous densities. The peptide conformation was confirmed by a randomized omit map procedure (29) (Fig. 1 B). A random error <0.25 Å was added to each of the refined coordinates that did not contain peptide. These coordinates with the peptide region omitted were subject to Refmac refinement, and an omit electron density map with coefficients 2fo-fc was calculated. The omit map clearly indicates the side chain orientations, although the density is broken for the main chain between peptide residues 4 and 5, as well as between residues 5 and 6 (Fig. 1 B). Fig. 1 C shows the final 2fo-fc map surrounding the peptide. The only significant densities (
3
level) in the fo-fc map is a small positive peak near residue P5. This, in combination with the relatively weak main chain density near residue P5, may indicate some flexibility around the middle of the peptide.
The coordinates of both complexes have been deposited with the Protein Data Bank (Brookhaven, NY). Identifiers are 1b0g and 1bz9 for A2/p1049 and Db/p1027, respectively.
|
Results and Discussion |
|---|
|
TopAbstractMaterials and MethodsResults and DiscussionReferences |
|---|
1/2 superdomain, the 3 domain, and β2m) are superimposed. There is a relative domain movement of the 3 domain and β2m with respect to the 1/2 superdomain when comparing Db/p1027 with Db/NP, as well as in comparing Db/p1027 with A2/p1049 (Fig. 2 A). This type of domain movement has been observed in other class I crystal structures (2), and it could be an effect of the packing differences in the different crystal forms of these class I/peptide complexes.
|
1/2 superdomain (all peptides in this paper are compared by superimposing the 1/ 2 superdomain), the ends of the peptide are essentially superimposable. However, the middle of the peptide (residues 3–7) has different backbone conformations and side chain orientations among these class I complexes (Fig. 2, B–D). This is consistent with earlier observations (3).
Peptide Binding in the Absence of a Canonical Anchor Residue.
Sequence analysis of the peptides eluted from Db complexes shows that Db strongly prefers Asn at P5 and has a certain degree of preference for Met, Ile, or Leu at P9 (30). These observations are consistent with the Db/NP crystal structure (19). Asn at P5 is deeply buried in a polar pocket defined by GluA9, GlnA70, GlnA97, and TyrA156 (Fig. 3 A), and forms hydrogen bonds with the last three residues. (Throughout this paper, we use A, B, and P to represent class I heavy chain, β2m, and peptide, respectively. GluA9 refers to residue 9 of heavy chain, which is a Glu.) MetP9 is buried in a hydrophobic pocket formed by residues PheA116, TryA123, IleA124, ThrA143, and TrpA147.
|
Db/p1027 is the only crystal structure determined to date of a class I with a peptide that does not have the normal anchor residues. What we have observed in the structure of Db/p1027 may be a general phenomenon. Other class I allotypes also occasionally bind peptides without primary anchor residues (31, 32). These peptides must also have compensated for the lost binding energy by using other residues within the peptide.
Comparison of TCR Contacting Regions in A2/p1049 and Db/p1027.
Several lines of evidence suggest that TCR docks on a common area and in a similar orientation on different class I/peptide complexes. In the four cocrystal structures of TCR/class I (6–9), the TCR is situated diagonally across the peptide-binding cleft with the TCR footprint centered on the peptide. The subunit of TCR is located close to the NH2 terminus of the peptide, while the β subunit is close to the COOH terminus of the peptide. Several contacts at the periphery of the TCR/class I interface involve conserved residues, which may help steer TCR into a general orientation. Recently, Smith and Lutz made exhaustive mutations on the 1 and 2 domains of class I molecule HLA-B7 and tested the recognition of these mutated molecules by 12 allospecific CTL clones. They concluded that the 12 CTL clones recognize a discrete surface with the bound peptide in the center. Although the boundaries of the TCR footprints from the 12 clones are not identical, they overlap largely.
Based on the crystal structure of A6/HLA-A2/Tax (6), the footprint of TCR can be approximated as a rectangular box with its width being 22 Å, measured from the C atom of A65 to A157. The length of the rectangular box is 32 Å, measured from the C atom of A170 to P9. The center of the rectangle is located near peptide residue P4. These dimensions are consistent with what is observed in the other TCR/MHC cocrystal structures (7, 8). It is also consistent with an earlier mutational study that suggests the TCR footprint is roughly 30 x 20 Å2 (34). This rectangular box contains the peptide and the majority of the 1/2 helices. In the following paragraphs, we first compare the peptide and the 1/2 helices in A2/p1049 and Db/p1027 at an atomic level. We then examine the shape and charge comparison of these two TCR footprints on the molecular surface diagrams.
The termini of p1027 and p1049 are fixed by a set of conserved hydrogen bonds and by specificity pockets that are complementary for the anchor residues, and thus they have similar conformations. The backbones of the first and last residues in p1049 and p1027 (Fig. 4 A) are essentially superimposable (rmsd 0.7 Å). The side chains of residues at the ends of the peptide (P1, P2, P8, and P9) have similar orientations. However, the average difference between C atoms of the middle residues (from P3 to P7) is 2.6 Å. The side chains of these middle residues also have different orientations (Fig. 4 A and Fig. 2 D). These differences result in different solvent accessibility patterns of the two peptides. In p1049, residue PheP5 is the most solvent-exposed residue (Fig. 4 B). Residues in p1027 are more solvent accessible than those in p1049, as seen by the much higher protrusion formed by ValP5. Residue TyrP8 in p1027 is also significantly exposed to solvent (Fig. 4 B).
|
1 and 2 helices were found to be in direct contact with TCR. These TCR-contacting residues do not completely overlap between the two crystal structures. Since we cannot predict which set of residues will be utilized by Db or A2 to interact with AHIII12.2, we will analyze all of these potential contacts. Based on the two cocrystal structures, there are 23 unique residues on A2/p1049 and Db/p1027 that may interact with the AHIII12.2 TCR. Interestingly, these 23 residues include important residues for all TCR/class I/peptide complexes examined to date (7). 10 of these 23 residues are different between A2 and Db (Table II and Fig. 5). Except for the conservative change from Ala to Gly at position A69, all the other changes involve dramatic charge, polarity, or size differences. Among the 13 residues with common amino-acid identities, some have taken different conformations between the two complexes (Fig. 5). Most of these common residues are highly conserved among different human or mouse class I alleles. For example, residue LysA66 is conserved among 23 different HLA-A allotypes, and the remaining 12 residues are conserved among at least 49 different HLA-A allotypes. These conserved residues are likely to help dock the TCR in a common orientation on class I/peptide and contribute to the binding energy between TCR and class I/peptide. However, the different residues between these complexes also have to contribute significantly to the binding energy in order to differentiate between different antigenic complexes.
|
|
|
P2 (Leu) in p1049 is buried in the HLA-A2 B pocket (35). P2 in p1027 points in the same direction, but is a much smaller residue (Ala). Neither makes any apparent contribution to the molecular surface in either structure. The position 3 residues (Trp in p1049 and Pro in p1027) are pointed into pocket D, away from the TCR contact surface. As can be seen in Fig. 3 C, these residues are almost completely buried and, similar to the P2 position, do not directly contribute to the molecular surface.
The next most prominent difference between the molecular surfaces is at the center of the peptide at P5. Peptide residue P5 protrudes higher in Db/p1027 than in A2/ p1049. The reason is obvious when comparing the orientations of P5 in p1027 and p1049 (Fig. 4 A and Fig. 2 D). ValP5 in p1027 points up, whereas PheP5 in p1049 mainly points sideways. This is also consistent with the solvent accessibility of position P5 in the two peptides (Fig. 4, B and C). The OE1 atom of GluA70 generates a negative charge above ValP5 in Db/p1027, but there is no surface charge in A2 because residue 70 is a His and is buried underneath PheP6.
Positions 6 and 7 are identical in composition (Phe and Pro, respectively, for both A2/p1049 and Db/p1027) but very different in orientation between the two complexes (Fig. 2 D and Fig. 4 A). Although both form hydrophobic patches, they constitute different topological surfaces to the right of the P5 protrusion. Position 8 is a Val in A2/p1049 and a Tyr in Db/p1027. These residues are both solvent exposed (Fig. 4 B), but their differing size and chemical nature results in different surface properties in the P8 region of each complex.
There are similarities between the two molecular surfaces at the COOH end of the peptide along the right-hand border of the TCR footprint, generated by a common residue LysA146. This highly conserved Lys is located above and forms a salt bridge with the carboxylate of the peptide and covers the COOH terminus. A hydrophobic area surrounds this positive charge at LysA146 in each complex. Close examination shows that the shapes of these hydrophobic areas are dissimilar. The depression observed beside the left edge of LysA146 in Fig. 6 is much shallower in Db/p1027 than in A2/p1049, due to three amino-acid changes. Residues ValP8, AlaA150, and AlaA149 in A2 are the much larger residues TyrP8, SerA150, and GlnA149 in Db. Additionally, the orientation of ProP7 is different between the two complexes and ProP7 contributes to the shallow depression in Db/p1027.
In summary, there appear to be few similarities in the TCR contact region of these two complexes when each area is examined in detail. There is similarity of gross features. The footprint is relatively acidic around P1, hydrophobic in the center, and basic around P9. However, these common features are largely formed by conserved residues in different class I allotypes. In A2/p1049, the acidic area around P1 is formed by residues GluA58, GluA61, and GluA63, and the positive charge around P9 is formed by residue LysA146. Residue A63 is conserved in 40 different HLA-A allotypes, and residues A58, A61, and A146 are conserved among 60 different HLA-A allotypes. The hydrophobic area in the center is largely formed by hydrophobic residues around peptide residue P5. Many peptides with hydrophobic residues at P5 and P6 do bind to A2 (30). Fig. 6 C shows the potential TCR contacting surface of a completely unrelated class I/peptide complex (A2/matrix peptide sequence GILGFVFTL) (3). It demonstrates that these common gross features exist on completely unrelated class I/peptide complexes. These features probably contribute to the binding of TCR in general and are most likely the features that result in the similar orientation of TCR on MHC (7, 8). However, they cannot be the sole contributions, as they would lead to the recognition of many unrelated A2/peptide complexes by this TCR, which is obviously not the case.
Potential Conformational Changes of the Peptide.
The A6/ HLA-A2/Tax cocrystal structure shows that the Tax peptide changes conformation upon binding by TCR (6). Comparisons of the molecular surfaces of the A2/p1049 and Db/p1027 complexes should also be evaluated with respect to potential conformational changes and to the new molecular surfaces that would be generated by such changes. Amino acid substitution experiments performed on A2/p1049 and Db/p1058 can provide us with some information about the potential conformational changes of peptide upon TCR binding. The only differences between p1027 (FAPGVFPYM) and p1058 (FAPGFFPYL) are two conservative changes at P5 and P9. Leu is a preferred omega anchor residue for Db bound peptides, but Met is a readily accepted substitute (30). The hydrophobic PheP5 in p1058 is not likely to bind in the polar pocket normally designed for AsnP5. It is more likely to be solvent exposed as is the Val in Db/p1027. Thus, we believe that p1058 binds to Db in a conformation roughly the same as p1027. The side chain orientations observed from the structure of Db/ p1027 are consistent with the TCR/MHC interaction data deduced from the amino acid substitution data generated previously (18). Therefore, it is likely that p1027 does not undergo any major conformational change upon TCR binding to Db/p1027.
The Tax peptide changed conformation in both TCR/ MHC cocrystal structures (6, 7). P6 moved away from the TCR and P7 moved toward the TCR. Residues P6 and P7 in p1049 may undergo similar conformational changes when engaging AHIII12.2. When ProP7 is substituted with Ala, CTL reactivity is reduced to 40% of the wild-type level (18). Yet, in the crystal structure, ProP7 points sideways toward the 2 helix and is solvent inaccessible. Thus, it is possible that ProP7 changes conformation and points up and contacts the TCR upon binding. Accompanying the change on P7, P6 may move down as observed in Tax. If this is the case, it will generate a deeper depression surrounding P5 which will make the shape of the hydrophobic region surrounding residue P5 (Fig. 6, A and D) similar to that in Db/p1027, but significant differences (especially charge distributions) would still exist between the footprints. Furthermore, although P8 in both peptides point up and are solvent accessible (Fig. 4, A and B), amino acid substitution experiments demonstrated that P8 in p1058 bound to Db is in contact with TCR, while P8 in p1049 bound to A2 is not (18). These data confirm that the interactions between AHIII12.2 TCR and the two complexes are different.
Mechanism of T Cell Allo- and Xeno-recognition.
The mechanism of T cell allo- and xeno-reactivity remains one of the paradoxes of modern immunology. Through positive and negative selection, T cells are educated in the thymus to recognize self-MHC complexed with foreign peptide in the periphery. The requirements for both MHC and peptide in selection are clear (36). However, even after thymic education, 1–10% of T cells still recognize allo- or xeno-MHC antigens that they could not have encountered before (14, 37). The most popular model proposed to answer this paradox is that the molecular surfaces of the allo- or xeno- and syngeneic MHC antigens are similar in shape or/and charge regardless of their derivation.
The comparison of the TCR contacting surfaces from the crystal structures shows that A2/p1049 and Db/p1027 are different. These observations suggest that TCR cross-recognition of these class I/peptide epitopes is not due to simple shape and/or charge mimicry. Model building experiments on different allo- or cross-reactive class I have reached similar conclusions (38, 39). These observations, combined with the amino acid substitution data (18), can be best interpreted using a functional mimicry model, where TCR makes contacts with both common and different features on the two class I/peptide complexes and the mimicry is functional instead of structural. The common features between the two complexes mostly come from conserved residues and may help to steer TCR into a common orientation on different class I/peptide complexes. However, the noncommon features must also contribute significantly to ensure the specificity of a TCR.
When an allo- or xenogeneic class I interacts with a TCR, some of the residues under the TCR footprint remain the same as in the syngeneic complex, but other residues, perhaps many, will be different between these complexes. Among those changed residues, some will lose their interaction with TCR, but others will make adventitious contacts with TCR. Presumably, the lost and gained interactions will come to a balance, and the overall binding affinity between different class I ligands and TCR will be sufficient to trigger activation of the T cell. The relative importance of MHC versus peptide in this interaction is not predicted. There have been reports of peptide-independent TCR recognition of allo-reactive class I (40). This would suggest, for that case at least, that the contacts between the allo-reactive class I and TCR are sufficient to trigger the T cell, regardless of the peptide bound. Note also that this model does not suggest that either the TCR or class I has to be locked into a specific conformation. Loops and side chains of TCR may move to best accommodate the common and different contacts on a particular class I/peptide complex.
Recently, Speir et al. (16) generated a model of an allogeneic complex (Ld/QL9), based on the crystal structure of Ld with a mixture of peptides, and compared the model with the crystal structure of a syngeneic murine class I/peptide complex (Kb/dEV8). Based on these studies, they hypothesized a modified molecular mimicry model, in which a critical local charge mimicry (formed by residues SerP7 and AspA77 in Kb/ dEV8 and AspP8 and AsnA77 in Ld/QL9) contributes to the recognition of these two complexes by the same TCR. The functional mimicry model proposed here does not rule out the possibility of this modified molecular mimicry, but we think the original and modified molecular mimicry model does not account for all TCR cross-reactivity.
The cocrystal structures of A2/Tax with two different TCRs (A6 and B7) further demonstrate the requirement for flexibility in our thoughts about TCR recognition. A6 and B7 recognize A2/Tax by a common docking mechanism, but a different set of residues on the TCR is used to interact with A2/Tax. Only 1 of the 17 residues on the B7 TCR that contact A2/Tax is also found in the A6/A2/Tax interface. An additional 12 TCR residues that contact A2/ Tax in the two TCRs are spatially equivalent, but they are different amino acids. Even when the TCR/MHC interactions were examined at an atomic level, few involved spatially equivalent and identical atoms. Most TCR/MHC interactions are between atomic pairs which are not spatially equivalent and/or have very different chemical natures. The sum of these common and different interactions must have generated sufficient affinity for the same class I to bind two different TCRs. It is not difficult to imagine that similar situations can happen between two different class I/peptide complexes with the same TCR.
Crystal structures of several antibody idiotope/antiidiotope complexes (41, 42) also demonstrate the absence of molecular mimicry between the antiidiotope and the antigen. Fields et al. (42) suggested that "the mimicking is functional, involving similar binding interactions, rather than exact topological replicas." For example, 6 out of the 12 hydrogen bonds are structurally (spatially) equivalent, although residues involved in these hydrogen bonds are amino acids with different size and chemical natures. We speculate that in many cases these interactions do not have to be structurally (or spatially) equivalent (as demonstrated by the cocrystal structure of TCRs A6 and B7 with A2/ Tax); however, sufficient binding energy between antibody or TCR with its cross-reactive ligands can still be generated.
The functional mimicry model implies that TCR cross-reactivity, like antibody cross-reactivity, is unpredictable. It is possible for cross-reactive ligands to be structurally dissimilar, yet functionally similar. This mechanism also explains why the TCR repertoire selected by a single peptide can still recognize dissimilar ligands (43, 44). The different ligands are functionally similar with respect to recognition by the TCR.
|
Acknowledgments |
|---|
Submitted: 29 July 1998
Revised: 4 November 1998
|
References |
|---|
|
TopAbstractMaterials and MethodsResults and DiscussionReferences |
|---|
1 Batalia MA & Collins EJ. Peptide binding by class I and class II MHC molecules, Biopolymers, 1997, 43, 281–302.[Medline]
2 Fremont DH, Matsumura M, Stura EA, Peterson PA & Wilson IA. Crystal structures of two viral peptides in complex with murine MHC class I H-2Kb, Science, 1992, 257, 919–927.
3 Madden DR, Garboczi DN & Wiley DC. The antigenic identity of peptide–MHC complexes: a comparison of the conformations of five viral peptides presented by HLA-A2, Cell, 1993, 75, 693–708.[Medline]
4 Garrett TPJ, Saper MA, Bjorkman PJ, Strominger JL & Wiley DC. Specificity pockets for the side chains of peptide antigens in HLA-AW68, Nature, 1989, 341, 692–696.[Medline]
5 Guo H-C, Jardetzky TS, Garrett TPJ, Lane WS, Strominger JL & Wiley DC. Different length polypeptides bind to HLA-Aw68 similarly at their ends but bulge out in the middle, Nature, 1992, 360, 364–366.[Medline]
6 Garboczi DN, Ghosh P, Utz U, Fan WR, Biddison WE & Wiley DC. Structure of the complex between human T-cell receptor, viral peptide and HLA-A2, Nature, 1996, 384, 134–141.[Medline]
7 Ding Y-H, Smith KJ, Garboczi DN, Utz U, Biddison WE & Wiley DC. Two human T cell receptors bind in a similar diagonal mode to the HLA-A2/Tax peptide complex using different TcR amino acids, Immunity, 1998, 8, 403–411.[Medline]
8 Garcia KC, Degano M, Pease LR, Huang M, Peterson PA, Teyton L & Wilson IA. Structural basis of plasticity in T cell receptor recognition of a self peptide– MHC antigen, Science, 1998, 279, 1166–1172.
9 Teng M-K, Smolyar A, Tse AGD, Liu J-H, Liu J, Hussey RE, Nathenson SG, Chang H-C, Reinherz EL & Wang J-H. Identification of a common docking topology with substantial variation among different TcR– peptide–MHC complexes, Curr Biol, 1998, 8, 409–412.[Medline]
10 Kersh GJ & Allen PM. Essential flexibility in the T-cell recognition of antigen, Nature, 1996, 380, 495–498.[Medline]
11 Grossman Z & Singer A. Tuning of activation thresholds explains flexibility in the selection and development of T cells in the thymus, Proc Natl Acad Sci USA, 1996, 93, 14747–14752.
12 Oldstone MBA. Molecular mimicry and autoimmune diseases, Cell, 1990, 50, 819–820.
13 Theofilopolous AN. The basis of autoimmunity. I. Mechanisms of aberrant self-recognition, Immunol Today, 1995, 16, 90–98.[Medline]
14 Matzinger P & Bevan MJ. Why do so many lymphocytes respond to major histocompatibility antigens? , Cell Immunol, 1977, 29, 1–5.[Medline]
15 Quaratino S, Thorpe CJ, Travers PJ & Londei M. Similar antigenic surfaces, rather than sequence homology, dictate T-cell epitope molecular mimicry, Proc Natl Acad Sci USA, 1995, 92, 10398–10402.
16 Speir JA, Garcia KC, Brunmark A, Degano M, Peterson PA, Teyton L & Wilson I. Structural basis of 2C TcR allorecognition of H-2Ldpeptide complexes, Immunity, 1998, 8, 553–562.[Medline]
17 Henderson RA, Cox AL, Sakaguchi K, Appella E, Shabanowitz J, Junt DF & Engelhard VH. Direct identification of an endogenous peptide recognized by multiple HLA-A2.1-specific cytotoxic T cells, Proc Natl Acad Sci USA, 1993, 90, 10275–10279.
18 Loftus DJ, Chen Y, Covell DG, Engelhard VH & Appella E. Differential contact of disparate class I/peptide complexes as the basis for epitope cross-recognition by a single TCR, J Immunol, 1997, 158, 3651–3658.[Abstract]
19 Young ACM, Zhang W, Sacchettini JC & Nathenson SG. The three-dimensional structure of H-2Dbat 2.4Å resolution: implications for antigen-determinant selection, Cell, 1994, 76, 39–50.[Medline]
20 Garboczi DN, Hung DT & Wiley DC. HLA-A2–peptide complexes: refolding and crystallization of molecules expressed in Escherichia coliand complexed with single antigenic peptides, Proc Natl Acad Sci USA, 1992, 89, 3429–3433.
21 Otwinowski, Z., and W. Minor. 1996. Processing of x-ray diffraction data collected in oscillation mode. In Methods in Enzymology. Vol. 276A. C.W.J. Carter and R.M. Sweet, editors. Academic Press, Inc., New York. 307–326.
22 Collaborative Computational Project Number 4. The CCP4 suite: programs for protein crystallography, Acta Crystal D, 1994, 50, 760–763.[Medline]
23 Jones TA, Zou J-Y, Cowan SW & Kjeldgaard M. Improved methods for building protein models in electron density maps and the location of errors in these models, Acta Crystal A, 1991, 47, 110–119.
24 Brunger, A. 1992. X-PLOR (Version 3.1) A System for X-Ray Crystallography and NMR. Yale University Press, New Haven, CT. 382 pp.
25 Murshudov GN, Vagin AA & Dodson EJ. Refinement of macromolecular structures by the maximum-likelihood method, Acta Crystal D, 1997, 53, 240–255.[Medline]
26 Brunger AT, Krukowski A & Erickson J. Slow-cooling protocols for crystallographic refinement by simulated annealing, Acta Crystal A, 1990, 46, 585–593.
27 Lamzin, V.S., and K.S. Wilson. 1997. Automated refinement for protein crystallography. In Methods in Enzymology. Vol. 277B. C.W.J. Carter and R.M. Sweet, editors. Academic Press, Inc., New York. 269–305.
28 Brunger AT. The free R value: a novel statistical quantity for assessing the accuracy of crystal structures, Nature, 1992, 355, 472–474.[Medline]
29 McRee, D.E. 1993. Practical Protein Crystallography. Academic Press, Inc., San Diego, CA. 386 pp.
30 Falk K, Rotzchke O, Stevanovic S, Jung G & Rammensee H-G. Allele-specific motifs revealed by sequencing of self-peptides eluted from MHC molecules, Nature, 1991, 351, 290–296.[Medline]
31 Mendoza LM, Paz P, Zuberi A, Christianson G, Roopenian D & Shastri N. Minors held by majors: the H13 minor histocompatibility locus defined as a peptide/ MHC class I complex, Immunity, 1997, 7, 461–472.[Medline]
32 Burrows GG, Ariail K, Celnik B, Gambee JE, Bebo BFJ, Offner H & Vandenbark AA. Variation in H-2K(k) peptide motif revealed by sequencing naturally processed peptides from T-cell hybridoma class I molecules, J Neuro Res, 1996, 45, 803–811.[Medline]
33 Smith KD & Lutz CT. Alloreactive T cell recognition of MHC class I molecules, J Immunol, 1997, 158, 2805–2812.[Abstract]
34 Ajitkumar P, Geier SS, Kesari KV, Borriello F, Nakagawa M, Bluestone JA, Saper MA, Wiley DC & Nathenson SG. Evidence that multiple residues on both the -helices of the class I MHC molecule are simultaneously recognized by the T cell receptor, Cell, 1988, 54, 47–56.[Medline]
35 Saper MA, Bjorkman PJ & Wiley DC. Refined structure of the human histocompatibility antigen HLA-A2 at 2.6Å resolution, J Mol Biol, 1991, 219, 277–319.[Medline]
36 Benoist C & Mathis D. Positive selection of T cells: fastidious or promiscuous? , Curr Opin Immunol, 1997, 9, 245–249.[Medline]
37 Kaye J & Janeway CA Jr. The Fab fragment of a directly activating monoclonal antibody that precipitates a disulfide-linked heterodimer from a helper T cell clone blocks activation by either allogeneic Ia or antigen and self-Ia, J Exp Med, 1984, 159, 1397–1412.
38 Brock R, Wiesmuller K-H, Jung G & Walden P. Molecular basis for the recognition of two structurally different major histocompatibility complex/peptide complexes by a single T-cell receptor, Proc Natl Acad Sci USA, 1996, 93, 13108–13113.
39 Tallquist MD, Weaver AJ & Pease LR. Degenerate recognition of alloantigenic peptides on a positive- selecting class I molecule, J Immunol, 1998, 160, 802–809.
40 Smith PA, Brunmark A, Jackson MR & Potter TA. Peptide-independent recognition by alloreactive cytotoxic T lymphocytes (CTL), J Exp Med, 1997, 185, 1023–1033.
41 Bentley GA, Boulot G, Riottot MM & Poljak RJ. Three-dimensional structure of an idiotope–anti-idiotope complex, Nature, 1990, 348, 254–257.[Medline]
42 Fields BA, Goldbaum FA, Ysern X, Poljak RJ & Mariuzza RA. Molecular basis of antigen mimicry by an anti-idiotope, Nature, 1995, 374, 739–742.[Medline]
43 Ignatowicz L, Kappler J & Marrack P. The repertoire of T cells shaped by a single MHC/peptide ligand, Cell, 1996, 84, 521–529.[Medline]
44 Liu CP, Parker D, Kappler J & Marrack P. Selection of antigen-specific T cells by a single IEk peptide combination, J Exp Med, 1997, 186, 1441–1450.
45 Nichols A, Sharp KA & Honig B. Protein folding and association: insights from the interfacial and thermodynamic properties of hydrocarbons, Proteins, 1991, 11, 281–296.[Medline]
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Facebook 
Reddit 
Technorati 
Twitter What's this?
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| TABLE OF CONTENTS |
|
