A Model for the Three-Dimensional Structure of Human Plasma Vitronectin from Small-Angle Scattering Measurements

Abstract

Small-angle X-ray scattering (SAXS) measurements were used to characterize vitronectin, a circulatory protein found in human plasma that functions in regulating cell adhesion and migration, as well as proteolytic cascades that affect blood coagulation, fibrinolysis, and pericellular proteolysis. SAXS measurements were taken over a 3-fold range of protein concentrations, yielding data that characterize a monodisperse system of particles with an average radius of gyration of 30.3 +/- 0.6 A and a maximum linear dimension of 110 A. Shape restoration was applied to the data to produce two models of the solution structure of the ligand-free protein. A low-resolution model of the protein was generated that indicates the protein to be roughly peanut-shaped. A better understanding of the domain structure of vitronectin resulted from low-resolution models developed from available high-resolution structures of the domains. These domains include the N-terminal domain that was determined experimentally by NMR [Mayasundari, A., Whittemore, N. A., Serpersu, E. H., and Peterson, C. B. (2004) J. Biol. Chem. 279, 29359-29366] and the docked structure of the central and C-terminal domains that were determined by computational threading [Xu, D., Baburaj, K., Peterson, C. B., and Xu, Y. (2001) Proteins: Struct., Funct., Genet. 44, 312-320]. This model provides an indication of the disposition of the central domain and C-terminal heparin-binding domains of vitronectin with respect to the N-terminal somatomedin B (SMB) domain. This model constructed from the available domain structures, which agrees with the low-resolution model produced from the SAXS data, shows the SMB domain well separated from the central and heparin-binding domains by a disordered linker (residues 54-130). Also, binding sites within the SMB domain are predicted to be well exposed to the surrounding solvent for ease of access to its various ligands.

A Deletion Mutant of Vitronectin Lacking the Somatomedin B Domain Exhibits Residual Plasminogen Activator Inhibitor-1-binding Activity

“…Fluorescence measurements on the complexes dem-onstrated energy transfer, and the FRET signal was concentration-dependent, so that increasing the ratio of vitronectin to PAI-1 decreased the FRET signal, in effect sequestering PAI-1 on separate vitronectin molecules by binding to the high affinity site in the somatomedin B domain. The FRET measurements indicate the binding sites to be separated by ϳ57 Å on the surface of vitronectin, which has an ellipsoid shape with a length of ϳ110 Å (14).…”

“…Small-angle x-ray scattering shows that the C-terminal domain of vitronectin lies within a separate lobe from the N-terminal somatomedin B domain (14). FRET was chosen as a reasonable approach to test the relative proximity of the PAI-1-binding sites within the bi-lobed structure.…”

“…Although there is some conflict among the structures that is rooted in the identification of disulfide bonds (6, 9 -13), in all of the structures on the somatomedin B domain, the most important feature is a single turn ␣-helix that defines the binding interface with PAI-1. Small-angle x-ray scattering on monomeric vitronectin generated a low resolution model that revealed a bi-lobed structure for vitronectin (14). Assembling the NMR and x-ray scattering data with the structure predicted from threading suggests that the C-terminal domain lies within a separate lobe from the N-terminal somatomedin B domain (14).…”

“…Small-angle x-ray scattering on monomeric vitronectin generated a low resolution model that revealed a bi-lobed structure for vitronectin (14). Assembling the NMR and x-ray scattering data with the structure predicted from threading suggests that the C-terminal domain lies within a separate lobe from the N-terminal somatomedin B domain (14).…”

Integrin-linked kinase is required for vitronectin-mediated internalization ofStreptococcus pneumoniaeby host cells

“…The impact of the heparin-binding site of the vitronectin molecule (Lynn et al, 2005;Xu et al, 2001) on vitronectin recruitment by pneumococci was investigated by using heparin as a competitor in binding experiments. Our flow cytometric analysis demonstrated a dose-dependent inhibition of vitronectin acquisition by heparin (Fig.…”