1. Immunology and Inflammation
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The structure of a LAIR1-containing human antibody reveals a novel mechanism of antigen recognition

  1. Fu-Lien Hsieh
  2. Matthew K Higgins  Is a corresponding author
  1. University of Oxford, United Kingdom
Short Report
Cite this article as: eLife 2017;6:e27311 doi: 10.7554/eLife.27311
3 figures and 2 tables

Figures

Figure 1 with 3 supplements
Structure of a LAIR1-containing antibody Fab fragment.

(A) The structure of the Fab fragment. LAIR1 (red) is inserted into the third CDR loop of the heavy chain (yellow) through two extended linkers (orange). The light chain is blue. The dashed orange link represents protein disordered in the structure. (B) The organization of the CDRs. The three CDR loops of the light chain and remaining two CDR loops of the heavy chain directly contact the LAIR1 insert or the linkers. Each of the CDR loops and its corresponding label is a shown in a different colour. (C) A disulphide bond between C93 of the light chain and C223 of the heavy chain stabilizes the interface (cysteine residues are shown as sticks).

https://doi.org/10.7554/eLife.27311.003
Figure 1—figure supplement 1
Annotated sequence of antibody MGD21 and its alignment to germ line LAIR1.

(A) The sequence of the light chain of MGD21 with the CDR loops indicated by blue bars. (B) The sequence of the heavy chain of MGD21 aligned to that of germ line LAIR1. Hexagons represent putative glycosyation sites with the red hexagon representing a site lost in MGD21. Yellow circles mark sites in germ line LAIR1 known to affect collagen binding. Red stars represent cysteine residues that make a cross-chain disulphide bond. Residues in the heavy chain are labeled according to whether they derive from the V, J or LAIR1 genes. In both cases, Kabat numbers (Dunbar and Deane, 2016) of residues are given above the sequence. In addition, all the CDRs, with the exception of the third CDR of the heavy chain, are labeled with their canonical class (Martin and Thornton, 1996).

https://doi.org/10.7554/eLife.27311.004
Figure 1—figure supplement 2
Electron density.

(A) A view of the electron density, showing the 2Fo-Fc map in blue, contoured at 1.0σ. (B) A view of the electron density in the region in which the CDR loops of the heavy chain (dark blue) contact the LAIR1 insert (red). (C) A view of the electron density where the CDR loops of the light chain (magenta) contact linker 2.

https://doi.org/10.7554/eLife.27311.005
Figure 1—figure supplement 3
Crystal packing and order.

(A) A superimposition of the two antibody molecules in the asymmetric unit, with chains A and B in yellow and C and D in red. (B) The structure of chains A and B shown in putty representation with putty thickness determined by B factor. The colour scale for blue to red is from B factors of 30 to 100.

https://doi.org/10.7554/eLife.27311.006
Structure and polymorphism in the LAIR1 insertion.

(A) An alignment of germ line LAIR1 (cyan) with the antibody LAIR1 insertion (red). (B) The residues that differ between the LAIR1 insertion in antibody MGD21 and germ line LAIR1 are shown as red sticks. (C) A surface representation of the structure of LAIR1 (grey) with residues whose mutation has a major (red) or minor (yellow) effect on collagen binding highlighted (Brondijk et al., 2010). (D) A surface view of the LAIR1 insert in antibody MGD21 (grey) with residues that differ from germ line LAIR1 highlighted (red). (E) A surface view of the LAIR1 insert (grey) with residues that differ from germ line LAIR1 in all 27 antibodies tested to date (Tan et al., 2016) highlighted (red). (F) A sequence alignment of germ line LAIR1 and the LAIR1 insert in the MGD21 antibody. Yellow circles are sites residues shown to play a role in collagen binding while a red hexagon represents a potential N-linked glycosylation site mutated in the LAIR1 insert.

https://doi.org/10.7554/eLife.27311.009
Comparison of the LAIR1-containing antibody with other unusual antibodies.

The structure of the LAIR1-containing monoclonal antibody is compared with a classical mouse monoclonal antibody (9AD4; PDB code 4U0R), a human monoclonal antibody with broadly neutralizing potential against HIV (PG16; PDB code 4DQ0) and a bovine monoclonal antibody (BLV5B8; PDB code 4K3E). In each case, the light chain is blue and the two immunoglobulin domains of the heavy chain are yellow. Inserted domains are shown in red with linker regions in orange.

https://doi.org/10.7554/eLife.27311.010

Tables

Table 1

Data collection and refinement statistics. The structure was determined from a single crystal. Values in parentheses are for highest-resolution shell. Rfree was determined using 1968 reflections (4.8%) The structure is deposited with pdb code 5NST.

https://doi.org/10.7554/eLife.27311.007

Fab-MGD21
Data collection
 Space groupC121
 Cell dimensions
a, b, c (Å)169.8, 86.5, 104.0
 α β γ (°)90.0, 126.7, 90.0
 Wavelength0.92819
 Resolution (Å)81.90–2.52 (2.56–2.52)
 Total Observations131833 (5451)
 Total Unique40946 (2031)
Rpim (%)5.4 (67.8)
Rmerge (%)8.3 (88.5)
Rmeas (%)9.9 (112.1)
CC1/20.992 (0.571)
I/σ(I)7.4 (1.0)
 Completeness (%)99.8 (98.3)
 Multiplicity3.2 (2.7)
 Wilson B factor55.216
Refinement
 Number of reflections40946
Rwork / Rfree21.9/26.7
 Number of residues
Protein1076
 R.m.s deviations
Bond lengths (Å)0.01
Bond angles (°)1.25
 All Atom clash score5
 B factors
All atoms71.53
Solvent63.12
Variable domains65.17
Constant domains74.29
LAIR1 insert73.70
Linkers94.71
 Ramachandran plot
Favored (%)95.2%
Allowed (%)4.8%
Disallowed (%)0.0%
Table 2

A list of interactions between the LAIR1 insert and linkers that occupies the heavy chain CDR3 loop and the other five CDR loops of the antibody.

https://doi.org/10.7554/eLife.27311.008
CDR loopResidueGroup LAIR1 regionResidueGroupInteraction
 Light chain CDR1Q27Side chainLinker 2A222Main ChainHydrogen Bond
 Light chain CDR2Y49Side chainLinker 1L102Side chainHydrophobic Packing
 Light chain CDR2N53Side chainLinker 1S104Side chainHydrogen Bond
 Light chain CDR3C93Side chainLinker 2C223Side chainDisulphide Bond
 Light chain CDR3F94Main ChainLinker 2E227Side ChainHydrogen Bond
 Heavy chain CDR1N32Side chainLAIR1R134Side chainHydrogen Bond
 Heavy chain CDR2R57Side ChainLAIR1P109Main ChainHydrogen Bond

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