PDBsum entry 1h5b

T cell receptor

PDB id: 1h5b

Contents

Protein chains

113 a.a. *

112 a.a. *

Ligands

GOL

Metals

_CL ×3

Waters ×215

* Residue conservation analysis

References listed in PDB file

Key reference

Title

Crystal structure of a t cell receptor valpha11 (av11s5) domain: new canonical forms for the first and second complementarity determining regions.

Authors

M.Machius, P.Cianga, J.Deisenhofer, E.S.Ward.

Ref.

J Mol Biol, 2001, 310, 689-698. [DOI no: 10.1006/jmbi.2001.4794]

PubMed id

11453680

Abstract

We describe the X-ray crystallographic structure of a murine T cell receptor (TCR) Valpha domain ("Valpha85.33"; AV11S5-AJ17) to 1.85 A resolution. The Valpha85.33 domain is derived from a TCR that recognizes a type II collagen peptide associated with the murine major histocompatibility complex (MHC) class II molecule, I-A(q). Valpha85.33 packs as a Valpha-Valpha homodimer with a highly symmetric monomer-monomer interface. The first and second complementarity determining regions (CDR1 and CDR2) of this Valpha are shorter than the CDRs corresponding to the majority of other Valpha gene families, and three-dimensional structures of CDRs of these lengths have not been described previously. The CDR1 and CDR2 therefore represent new canonical forms that could serve as templates for AV11 family members. CDR3 of the Valpha85.33 domain is highly flexible and this is consistent with plasticity of this region of the TCR. The fourth hypervariable loop (HV4alpha) of AV11 and AV10 family members is one residue longer than that of other HV4alpha regions and shows a high degree of flexibility. The increase in length results in a distinct disposition of the conserved residue Lys68, which has been shown in other studies to play a role in antigen recognition. The X-ray structure of Valpha85.33 extends the database of canonical forms for CDR1 and CDR2, and has implications for antigen recognition by TCRs that contain related Valpha domains.

Figure 1.
Figure 1. Stereo Figure of the structure of Va85.33. (a) 2F[o] - F[c] omit map in the region around CDR1. (b) Superposition of the C^a backbone traces of the two Va85.33 dimers in the asymmetric unit. The backbone is colored according to atomic displacement parameters (blue = 18 Å2, RED = 100 Å2). Superpositions were done with the program SPDBViewer.[50] The complementarity determining regions, the fourth hypervariable region (HV4) as well as the C and N termini are indicated. All Figures were generated with Bobscript, [51] gl_render (L. Esser, University of Texas Southwestern Medical Center, unpublished) and PovRay (Persistence of Vision Ray Tracer, v3.02, POV-Team, www.povray.org). Methods: the Va85.33 domain containing a His[6]-tag was expressed as a secreted protein in Escherichia coli as described.[35] Crystals were obtained at 20 °C by vapor diffusion from drops containing 3 µl of protein (5 mg ml -1 in 50 mM Tris-HCl (pH 8.0), 100 mM NaCl) plus 3 µl of reservoir solution (100 mM sodium citrate-HCl, 1.4-1.7 M lithium chloride, pH 5.0-6.0) equilibrated against 1 ml of reservoir solution. Hexagonal crystals appeared after three to ten days and grew to a final size of 0.7 mm diameter and 0.3 mm thickness within one to three weeks. Va85.33 crystallized with the symmetry of space group P3[2]21 with cell constants of a = b = 83.5 Å, c = 132.1 Å, and four molecules per asymmetric unit. Prior to data collection, the crystals were cryo-protected by transferring them into harvesting solution (100 mM sodium citrate-HCl, 2 M lithium chloride, pH 5.5) supplemented with up to 40 % (v/v) glycerol and flash-cooled in liquid propane. The crystals diffracted to 1.85 Å Bragg spacing when using synchrotron radiation. The structure was solved by multiple anomalous dispersion (MAD) using a seleno-methionine variant (two methionine residues per molecule). The seleno-methionine variant of Va85.33 was expressed in the methionine-auxotroph E. coli strain B834 grown in minimal medium supplemented with the natural amino acids and seleno-methionine. Purification and crystallization behavior was essentially unchanged compared to native Va85.33. The MAD experiment was carried out at beamline 19-ID (SBC-CAT) at the Advanced Photon Source (Argonne National Laboratory, Argonne, Illinois, USA). Data were indexed, integrated and scaled with the HKL2000 program package.[52] Eight selenium sites were identified at 2.5 Å resolution by direct methods (Shake'n'Bake 2.0[53]) using the data set collected at the energy for the selenium absorption peak. Selenium parameters were refined and the resulting phases (figure of merit 0.49) were improved by density modification using programs from the CCP4 package, [54] resulting in a figure of merit of 0.69. Model building was done with the program O. [55] Structure refinement was carried out with the program CNS v0.5 [56] employing cycles of simulated annealing, conjugate gradient minimization and calculation of individual atomic displacement parameters. Calculation of overall anisotropic displacement parameters and bulk solvent correction was used throughout. No non-crystallographic symmetry restraints were used at the highest resolution (1.85 Å). Water molecules were added where stereochemically reasonable after the protein part was completed. Side-chains with poorly defined density were truncated to alanine for refinement purposes. The final model contains residues 2 to 112 for molecule 1, residues 2 to 110 for molecule 2, residues 2 to 112 for molecule 3, residues 2 to 110 for molecule 4, three chloride ions, three glycerol molecules and 266 water molecules. The correctness of the model was confirmed through simulated annealing omit maps. The R[free] value is 23.6 % and the R[work] value is 21.9 % (Table 1).

Figure 3.
Figure 3. Stereo figures of the complementarity determining regions (CDRs) and the fourth hypervariable region (HV4). (a) CDR1 of Va85.33 (carbon atoms are shown in gray) superimposed on the CDR1 of A6 (carbon atoms are shown in cyan). (b) CDR2 of Va85.33 (carbon atoms are shown in gray) superimposed on the CDR1 of A6 (carbon atoms are shown in cyan). (c) The backbone of the Va85.33-CDR3 regions of all four molecules in the asymmetric unit (colored in red, green, blue and yellow) superimposed on the CDR3 regions of A6, TCR, B.4.2.3, KB5, N15-C20, B7, BM3.3, HA1.7, and 1934.4 (colored in gray). (d) The backbone of the Va85.33-HV4 regions of all four molecules in the asymmetric unit (colored in red, green, blue and yellow) superimposed on the HV4 regions of A6, TCR, B.4.2.3, KB5, N15, B7, D10, BM3.3, HA1.7, and 1934.4 (colored in gray).

The above figures are reprinted by permission from Elsevier: J Mol Biol (2001, 310, 689-698) copyright 2001.

Secondary reference #1

Title

Canonical structures for the hypervariable regions of t cell alphabeta receptors.

Authors

B.Al-Lazikani, A.M.Lesk, C.Chothia.

Ref.

J Mol Biol, 2000, 295, 979-995. [DOI no: 10.1006/jmbi.1999.3358]

PubMed id

10656805

Figure 1.
Figure 1. A schematic drawing of a T cell αβ receptor and its binding site. On the left we show the arrangements of the V and C domains and the helices that attach the receptor to the cell surface. On the right we show a schematic drawing of the T cell binding site. The hypervariable regions are labelled α1, α2, α3, β1, β2 and β3. They are attached to strands of β-sheet that are approximately perpendicular to the page and are shown as open squares.

Figure 6.
Figure 6. Schematic drawings of the size and main-chain hydrogen bonds of the second hypervariable regions in the Vδ of ES204 and for the canonical structures that form the second hypervariable region in currently known Vβ, V[H] and V[L] structures. Details of the Vβ conformations are given in Figure 5 and Table 2. Details of the V[H] and V[L] conformations are given by [Al-Lazikani et al 1997]. The region unique to each canonical structure is enclosed by a box.

The above figures are reproduced from the cited reference with permission from Elsevier