FC0100
Osteopontin (OPN)  -  Heparin

Biological function
Osteopontin is an extracellular matrix protein (sialoprotein) that functions in cell attachment by displaying a RGD cell adhesion sequence and as a cytokine that signals through integrin and CD44 cell adhesion molecules. OPN is also implicated in human tumor progression and cell invasion. OPN has intrinsic transforming activity, and elevated OPN levels promote metastasis.

Structural evidence
The PRE data indicate that the central segment (residues 90–150) of OPN expands upon heparin binding, comparable to an “unfolding-upon-binding” event. ¹⁵N relaxation measurements show that residues 90–120 exhibit increased backbone flexibility as evidenced by both decreased R₂ values and more negative heteronuclear NOEs consistent with higher conformational flexibility in the bound state. NMR measurements show that hydrodynamic radius of OPN increases through heparin binding from 3.6 nm to 5.2 nm. Double electron–electron resonance (DEER) measurements on six spin-labeled Cys double mutants indicating increased distances between the labeling sites of the six double mutants, in accord with the expansion of OPN upon heparin binding.

Biochemical evidence
Based on ITC measurements the ΔH and ΔS values for OPN-heparin binding are -16.3 kcal mol^-1 and -35 cal mol ^-1K^-1, giving a dissociation constant of 34 μM. A positive differential heat capacity, ΔCp, (about 40 calmol^-1K^-1) for heparin binding further supports that OPN in the bound state exhibits more conformational flexibility than in the free state.

Structure/Mechanism
The large negatively charged patch around residues 80–120 is likely to experience electrostatic repulsion from heparin and is, therefore, dis- placed from the central region of OPN around residue 155 upon heparin binding. OPN–heparin complex reveals an electrostatic binding mode largely governed by complementary charge patterns.

Mechanism category
tethering

Significance
Fuzziness provides compensation for the entropic penalty of binding and contributes to modulating the lifetime and rates of conversions of individual bound states.

Submitted by
Robert Konrat   Robert.Konrat@univie.ac.at