Architecture effects on L-selectin shedding induced by polypeptide-based multivalent ligands

Abstract

Multivalent interactions between selectins and their ligands play key roles in mediating the rolling and tethering of leukocytes in the early steps of the inflammatory response, as well as in lymphocyte circulation. L-selectin shedding, which is the proteolytic cleavage of L-selectin, can be induced by L-selectin clustering through the binding of multivalent ligands to multiple L-selectin molecules, and it has been shown to regulate leukocyte rolling and subsequent integrin activation for firm adhesion. In this paper, we report the production of homogenous glycopolypeptides modified with a 3,6-disulfo-galactopyranoside equipped with a caproyl linker. The saccharide residue was chemically attached to various polypeptide backbones of differing architectures; the composition and purity of the sulfated glycopolypeptides was confirmed via 1H-NMR spectroscopy, amino acid analysis (AAA), and electrophoretic analysis. The retention of the conformation of the polypeptide backbone was confirmed via circular dichroic spectroscopy. The shedding of l-selectin from the surface of Jurkat cells induced by these sulfated glycopolypeptides, determined via ELISA-based methods, varied based on differences in the architectures of the polypeptide scaffolds, suggesting opportunities for these strategies in probing cell-surface receptor arrays and directing cell signaling events.

L-selectin shedding is activated specifically within transmigrating pseudopods of monocytes to regulate cell polarity in vitro

“…2). Ligand density is known to act as a potent driver of shedding (43)(44)(45), and an increasing gradient of HSPG-like ligands across postcapillary venules could therefore influence L-selectin shedding. Also, a lack of shear stress in the subendothelial space could dramatically influence L-selectin clustering and subsequent signaling events that could feedback to activate ADAM17 locally and drive the shedding response specifically in pseudopods.…”

Glycopolymer probes of signal transduction

“…148 Using a polypeptide backbone, sialylated ligands were appended to give a polymer with ligands separated by distances estimated to range from 17 – 35 Å and a polymer with carbohydrate moieties separated by 35 – 50 Å. The polymer with shorter carbohydrate-spacing was more effective at eliciting L-selectin shedding than the polymer with the longer spacing.…”

Facile Supermolecular Aptamer Inhibitors of L-Selectin

“…It should also be noted that studies of L-selectin monovalent and bivalent aptamers have not demonstrated shedding [ 4 – 6 ]. Interestingly, synthetic multivalent carbohydrate ligands only induced shedding of L-selectin at millimolar concentrations while blocking L-selectin function at micromolar concentrations [ 9 ]. It has been hypothesized that shedding of L-selectin by multivalent ligands depends on both the number of ligands as well as the distance between the ligands [ 9 ].…”

“…Interestingly, synthetic multivalent carbohydrate ligands only induced shedding of L-selectin at millimolar concentrations while blocking L-selectin function at micromolar concentrations [ 9 ]. It has been hypothesized that shedding of L-selectin by multivalent ligands depends on both the number of ligands as well as the distance between the ligands [ 9 ]. In the future, we plan to tailor the parameters of the LS-Multi-Aptamer to directly determine if the physical properties of the Multi-Aptamers can be optimized to mediate shedding versus blocking.…”