Structural insights into fibronectin type III domain-mediated signaling

Abstract

The alternatively spliced type III extradomain B (EIIIB) of fibronectin (FN) is expressed only during embryogenesis, wound healing and tumorigenesis. The biological function of this domain is unclear. We describe here the first crystal structure of the interface between alternatively spliced EIIIB and its adjacent FN type III domain 8 (FN B-8). The opened CC' loop of EIIIB, and the rotation and tilt of EIIIB allow good access to the FG loop of FN-8, which is normally hindered by the CC' loop of FN-7. In addition, the AGEGIP sequence of the CC'' loop of EIIIB replaces the NGQQGN sequence of the CC' loop of FN-7. Finally, the CC'' loop of EIIIB forms an acidic groove with FN-8. These structural findings warrant future studies directed at identifying potential binding partners for FN B-8 interface, linking EIIIB to skeletal and cartilaginous development, wound healing, and tumorigenesis, respectively.

Figure 1. Crystal structure of FN-B8

(A) The primary and secondary structures of type III fibronectin domain 7, B and 8 are shown in green, black and magenta, respectively. The sequences of CC′ loops of FN-7, FN-B and FN-8 are highlighted. (B) A molscript diagram of FN B-8 structure. The β sandwiches are shown in yellow and red. (C) 2.01 Å simulated annealing omit fofc electron density map contouring at 3.0σ. The map was calculated via |Fobs| − |Fcalc|, ϕcalc, with these amino acids and a sphere 1.0 Å around them omitted prior to annealing and map calculation. Part of β-C (residues 125–127), β-F (residues 165–167) and β-G (residues 175–177) strands of EIII-B are shown here. (D) Superimposition of our EIII-B structure with previously solved NMR structure (RSCB code: 2fnb) demonstrates the similar overall folds.

Figure 1. Crystal structure of FN-B8

(A) The primary and secondary structures of type III fibronectin domain 7, B and 8 are shown in green, black and magenta, respectively. The sequences of CC′ loops of FN-7, FN-B and FN-8 are highlighted. (B) A molscript diagram of FN B-8 structure. The β sandwiches are shown in yellow and red. (C) 2.01 Å simulated annealing omit fofc electron density map contouring at 3.0σ. The map was calculated via |Fobs| − |Fcalc|, ϕcalc, with these amino acids and a sphere 1.0 Å around them omitted prior to annealing and map calculation. Part of β-C (residues 125–127), β-F (residues 165–167) and β-G (residues 175–177) strands of EIII-B are shown here. (D) Superimposition of our EIII-B structure with previously solved NMR structure (RSCB code: 2fnb) demonstrates the similar overall folds.

Figure 1. Crystal structure of FN-B8

(A) The primary and secondary structures of type III fibronectin domain 7, B and 8 are shown in green, black and magenta, respectively. The sequences of CC′ loops of FN-7, FN-B and FN-8 are highlighted. (B) A molscript diagram of FN B-8 structure. The β sandwiches are shown in yellow and red. (C) 2.01 Å simulated annealing omit fofc electron density map contouring at 3.0σ. The map was calculated via |Fobs| − |Fcalc|, ϕcalc, with these amino acids and a sphere 1.0 Å around them omitted prior to annealing and map calculation. Part of β-C (residues 125–127), β-F (residues 165–167) and β-G (residues 175–177) strands of EIII-B are shown here. (D) Superimposition of our EIII-B structure with previously solved NMR structure (RSCB code: 2fnb) demonstrates the similar overall folds.

Figure 1. Crystal structure of FN-B8

(A) The primary and secondary structures of type III fibronectin domain 7, B and 8 are shown in green, black and magenta, respectively. The sequences of CC′ loops of FN-7, FN-B and FN-8 are highlighted. (B) A molscript diagram of FN B-8 structure. The β sandwiches are shown in yellow and red. (C) 2.01 Å simulated annealing omit fofc electron density map contouring at 3.0σ. The map was calculated via |Fobs| − |Fcalc|, ϕcalc, with these amino acids and a sphere 1.0 Å around them omitted prior to annealing and map calculation. Part of β-C (residues 125–127), β-F (residues 165–167) and β-G (residues 175–177) strands of EIII-B are shown here. (D) Superimposition of our EIII-B structure with previously solved NMR structure (RSCB code: 2fnb) demonstrates the similar overall folds.

Figure 2. EIIIB alters the fibronectin structure

(A) Superimposition of EIIIB-8 of FN B-8 structure (shown in yellow/red) with FN 7–8 previously solved, RSBC code: 1fnf (shown in green), demonstrates that the differences in torsion and bending angles of FN B-8 compared with the FN 7–8; and the conformational differences of the AB, EF, and CC′ loops in EIIIB and FN-7. The two orientations are related by a rotation of 180°. (B) Molecular electrostatic potential surface representation of FN B-8 is colored by charge distribution (red negative, white neutral and blue positive) using program Pymol (DeLano, W.L., www.pymol.org).

Figure 2. EIIIB alters the fibronectin structure

(A) Superimposition of EIIIB-8 of FN B-8 structure (shown in yellow/red) with FN 7–8 previously solved, RSBC code: 1fnf (shown in green), demonstrates that the differences in torsion and bending angles of FN B-8 compared with the FN 7–8; and the conformational differences of the AB, EF, and CC′ loops in EIIIB and FN-7. The two orientations are related by a rotation of 180°. (B) Molecular electrostatic potential surface representation of FN B-8 is colored by charge distribution (red negative, white neutral and blue positive) using program Pymol (DeLano, W.L., www.pymol.org).