Structure determination of an anti-HIV-1 Fab 447-52D-peptide complex from an epitaxially twinned data set - PubMed (original) (raw)

Structure determination of an anti-HIV-1 Fab 447-52D-peptide complex from an epitaxially twinned data set

Amandeep K Dhillon et al. Acta Crystallogr D Biol Crystallogr. 2008 Jul.

Abstract

Although antibodies against the third variable loop (V3) of the HIV-1 viral envelope glycoprotein are among the first neutralizing antibodies to be detected in infected individuals, they are normally restricted in their specificity. X-ray crystallographic studies of V3-specific antibodies have contributed to a more thorough understanding of recognition of this epitope and of conserved features in the V3 loop that could potentially aid in the design of a multi-component vaccine. The human antibody 447-52D exhibits relatively broad neutralization of primary viral isolates compared with other V3-loop antibodies. A crystal structure of Fab 447-52D in complex with a V3 peptide (UG1033) was determined at 2.1 angstroms resolution. The structure was determined using an epitaxially twinned data set and in-house programs to detect and remove overlapping reflections. Although the processed data have lower than desired completeness and slightly higher than normal R values for the resolution, good-quality electron-density maps were obtained that enabled structure determination. The structure revealed an extended CDR H3 loop that forms a beta-sheet with the peptide, with the predominant contacts being main-chain hydrogen bonds. The V3 peptide and Fab show high structural homology with the previously reported structures of other Fab 447-52D complexes, reinforcing the idea that the V3 loop may adopt a small set of conserved structures, particularly around the crown of the beta-hairpin.

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Figures

Figure 1

Raw data from the epitaxially twinned data set. The frames are displayed with the XDISPLAYF (Otwinowski & Minor, 1997 ▶) software, and the images have been cropped to show only the strongly diffracting spots. (a) The first diffraction image (crystal to detector distance = 250 cm, oscillation angle = 0.25°). (b) Spots (diffraction maxima) corresponding to the first lattice are shown in green, after separation of the spot file into two files for the two different lattices, as described in §3. (c) Spots (green) corresponding to the second lattice. (d) The same image with all spots (green) from both lattices selected for auto-indexing by XDISPLAYF.

Figure 2

Stereoview of the Fab 447-52D–UG1033 peptide complex. The Fab is colored gray, with the light chain oriented to the left and the heavy chain oriented to the right. Complementarity-determining regions (CDRs) L1, L2 and L3 are colored purple, orange and cyan, respectively. CDRs H1, H2 and H3 are colored blue, green and red, respectively. The UG1033 V3 peptide is colored yellow. This and subsequent figures were generated with PyMOL (DeLano, 2002 ▶).

Figure 3

Stereoview of electron density of the UG1033 peptide bound to Fab 447-52D. The 2_F_ o − F c electron density (gray) for the UG1033 V3 peptide (yellow) after refinement is contoured at 1.0σ.

Figure 4

Binding-site interactions between the UG1033 V3 peptide and Fab 447-52D. The Fab CDR H3 is shown in violet and the UG1033 peptide in yellow. (a) The mixed three-strand β-sheet formed by CDR H3 and the V3 peptide is shown. (b) The main-chain hydrogen-bond interactions in the three-strand β-sheet are represented by black dotted lines. (c) Side-chain interactions between the UG1033 peptide and the Fab light chain (cyan) and heavy chain (violet).

Figure 5

Comparison of the UG1033 and MN peptides bound to Fab 447-52D. A superposition of the Cα backbones of the UG1033 V3 peptide bound to Fab 447-52D and the MN V3-peptide structure previously solved in complex with Fab 447-52D is shown. The UG1033 peptide is shown in yellow and the MN peptide in blue. The UG1033-peptide residues are labeled.

Figure 6

Comparison of V3 crystal structures. The Cα-backbone structures are shown in a similar orientations for (a) the UG1033 V3 peptide bound to Fab 447-52D (b) the UG1033 V3 peptide bound to Fab 2219 (PDB code

2b1a

) and the V3 loops from two gp120 core complexes: (c) PDB code

2b4c

and (d) PDB code

2qad

. Two conformations were observed for the V3 crown in the core gp120 V3 structure with PDB code

2b4c

and both of these conformations are shown in (c).

Figure 7

Comparison of the crystal structure of the UG1033 peptide bound to Fab 447-52D with the NMR structures of the MN and IIIB peptides bound to 447-52D Fv. The Cα-backbone structures are shown in a similar orientation for (a) the UG1033 V3 peptide from the crystal structure and (b) MN (PDB code

1n1z

) and (c) IIIB (PDB code

1u6u

) from NMR structures. The N-terminus of each peptide is oriented to the left.

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Structure determination of an anti-HIV-1 Fab 447-52D-peptide complex from an epitaxially twinned data set - PubMed (original) (raw)