A randomly coiled, high-molecular-weight polypeptide exhibits increased paracellular diffusion in vitro and in situ relative to the highly ordered alpha-helix conformer

Abstract

Purpose: The current investigation was conducted to examine the effect of secondary structure of model polypeptides on their hindered paracellular diffusion.

Methods: Poly-D-glutamic acid (PDGlu) was selected as one of the model polypeptides because of its ability to form two secondary structures; a negatively charged random coil and an alpha-helix with partial negative charge at pH 7.4 and 4.7, respectively. Poly-D-lysine (PDL) was selected as a positively charged random coil conformation at pH 7.4. Transport experiments were conducted across both a Caco-2 cell monolayer and the intestinal membrane of Sprague-Dawley rats. Additionally, using NMR, an estimation for the diffusion coefficient and the equivalent hydrodynamic radius for each model polypeptide was obtained.

Results: PDGlu in the randomly coiled conformation exhibited greater paracellular transport when compared to either the same polypeptide having an alpha-helix secondary structure or the positively charged, randomly coiled PDL.

Conclusions: Randomly coiled PDGlu was able to permeate through the negatively charged tight junctions of both biological membranes to a greater extent than PDGlu having an alpha-helix structure and suggests that molecular flexibility associated with the random coil conformation may play a more important role than overall charge and hydrodynamic radius on its hindered paracellular diffusion.