Application of recent advances in hydrodynamic methods for characterising mucins in solution

Mucins are the primary macromolecular component of mucus-nature's natural lubricant-although they are poorly characterised heterogeneous substances. Recent advances in hydrodynamic methodology now offer the opportunity for gaining a better understanding of their solution properties. In this study a combination of such methods was used to provide increased understanding of a preparation of porcine intestinal mucin (PIM), MUC2 mucin, in terms of both heterogeneity and quantification of conformational flexibility. The new sedimentation equilibrium algorithm SEDFIT-MSTAR is applied to yield a weight average (over the whole distribution) molar mass of 7.1 x 10(6) g mol(-1), in complete agreement with size exclusion chromatography coupled with multi-angle light scattering (SEC-MALS), which yielded a value of 7.2 x 10(6) g mol(-1). Sedimentation velocity profiles show mucin to be very polydisperse, with a broad molar mass distribution obtained using the Extended Fujita algorithm, consistent with the elution profiles from SEC-MALS. On-line differential pressure viscometry coupled to the SEC-MALS was used to obtain the intrinsic viscosity [eta] as a function of molar mass. These data combined with sedimentation coefficient data into the global conformation algorithm HYDFIT show that PIM has a flexible linear structure, with persistence length L (p) similar to 10 nm and mass per unit length, M (L) similar to 2380 g mol(-1) nm(-1), consistent with a Wales-van Holde ratio of similar to 1.2 obtained from the concentration dependence of the sedimentation coefficient.