Conformation of heparin studied with macromolecular hydrodynamic methods and X-ray scattering

Abstract

The hydrodynamic characteristics of heparin fractions in a 0.2 M NaCl solution have been determined. Experimental values varied over the following ranges: the sedimentation coefficient (at 20.0 degrees C), 1.3<s0x10(13)<3.2 s; the Gralen coefficient (sedimentation concentration-dependence parameter), 10<ks<70 cm3 g(-1); the translational diffusion coefficient, 3.9< D0x10(7)<15.4 cm2 s(-1); the intrinsic viscosity, 7.9<[eta]<40 cm3 g(-1). Combination of s0 with D0 using the Svedberg equation yielded molecular weights in the range 3.9<Mx10(-3)<37 g mol(-1). The value of the mass per unit length of the heparin molecule, ML, was determined using the theory of hydrodynamic properties of a weakly bending rod, giving ML=570 +/- 50 g nm(-1) mol(-1). The equilibrium rigidity, Kuhn segment length (A=9 +/- 2 nm) and hydrodynamic diameter (d=0.9 +/- 0.1 nm) of heparin were evaluated on the basis of the worm-like coil theory without the excluded volume effect, using the combination of hydrodynamic data obtained from fractions of different sizes. Small-angle X-ray scattering for three heparin fractions allowed an estimate for the cross-sectional radius of gyration as 0.43 nm; from the evolution with the macromolecule contour length of the radius of gyration, a value for the Kuhn segment length of 9 +/- 1 nm was obtained. A good correlation is thus observed for the conformational parameters of heparin from hydrodynamic and X-ray scattering data. These values describe heparin as a semi-rigid polymer, with an equilibrium rigidity that is essentially determined by a structural component, the electrostatic contribution being negligible in 0.2 M NaCl.

Quantitative Characterization of Heparin Binding to Tau Protein

“…The specific refractive increment of heparin (B in equations below) was determined experimentally. A calibration curve of refractive index versus heparin concentration was constructed, and the best fit value of the slope dñ/dw B ϭ 0.126 cm 3 /g is in agreement with the published values (23,24). A composition gradient was created using the Calypso system (Wyatt, Santa Barbara, CA), in which the fraction of solution A (f A ) is decreased from 1 to 0 in steps of 0.08, and the total concentrations of Tau (A) and heparin (B) in each of these mixtures is given by the following equations.…”

Pharmacology of Heparin and Related Drugs

“…In the course of the NMR conformational studies, it became clear that heparin did not behave in this way (Forster et al, 1989;Mulloy et al, 1993); the quantitative nuclear Overhauser effect values could not be interpreted unless it was assumed that heparin tumbled as an "asymmetric top," specifically as if the shape of the heparin molecules was more like a cylinder than a sphere. Analytical ultracentrifugation and X-ray scattering results were consistent in this assessment (Pavlov et al, 2003) and indicated a semirigid cylindrical structure for heparin oligosaccharides, with flexibility increasing as oligosaccharide length increases .…”

Binding between the Integrin αXβ2 (CD11c/CD18) and Heparin

“…Thus, it appears that the heparin binding by the ␣X I domain is a rare case of a Mg 2ϩ ion contributing to protein-heparin interaction. The end-to-end length of heparin either determined from hydrodynamic measurements (25) or direct measurements on heparin in complex with thrombin resolved by x-ray crystallography (26) corresponds to 0.5 nm/monosaccharide unit. dp12 oligomers would thus be assumed to take a length of 6 nm, which is comparable with the diameter of the I domain at 5 nm (12).…”