Versatile Methodology for Glycosurfaces: Direct Ligation of Nonderivatized Reducing Saccharides to Poly(pentafluorophenyl acrylate) Grafted Surfaces via Hydrazide Conjugation

Abstract

In this work, we report a convenient and versatile strategy for surface-grafted glycopolymer constructs with the goal of surface modification that controls the chemical presentation and grafting density of carbohydrate side chains. This approach employs a difunctional hydrazine linker, chemically modified to an active ester containing poly(pentafluorophenyl acrylate) grafted scaffold, to conjugate a variety of saccharides through the reducing end. The successive conjugation steps are carried out under mild conditions and yield high surface densities of sugars, as high as 4.8 mnol·cm−2, capable of multivalency, with an intact structure and retained bioactivity. We also demonstrate that this glycosylated surface can bind specific lectins according to the structure of its pendant carbohydrate. To demonstrate bioactivity, this surface platform is used to study the binding events of a human respiratory tract pathogen, Mycoplasma pneumoniae, on surfaces conjugated with sialylated sugars.

Sialylated Receptor Setting Influences Mycoplasma pneumoniae Attachment and Gliding Motility

“…Here we sought instead to examine mycoplasma binding and gliding activity on surfaces having well defined oligosaccharide presentation. We recently described the use of hydrazide conjugation to ligate reducing sugars to poly(PFPA) (pentafluorophenylacrylate)‐grafted surfaces (Chen et al, ). By varying the ratio of hydrazine and ethanolamine in the conjugation, it is possible to control the density of hydrazide available for ligating α‐2,3‐ and α‐2,6‐sialyllactose to the poly(PFPA) scaffold (Fig.…”

“…). We assessed the chemical modifications at each step by ellipsometry, drop shape analysis and Fourier transform infrared spectroscopy for thickness, hydrophobicity and chemical changes of the thin polymer film, respectively (Chen et al, ; data not shown). On average, ~10 nm hydrophilic glycosurfaces were built on ~15‐nm poly(PFPA) hydrophobic films.…”

“…This prompted us to pursue an approach that overcomes these limitations and yields functionalized surfaces with more precisely defined oligosaccharide presentations. To this end we recently described a methodology for ligating oligosaccharides at the reducing end to poly(PFPA)-grafted surfaces through a hydrazide linkage (Chen et al, 2017). Here we expanded upon that approach, varying the density of hydrazide linkages available by changing the ratios of hydrazine and competing ethanolamine in the conjugation to poly(PFPA) (Fig.…”

“…All glass coverslip substrates with grafted poly(PFPA) brushes were fabricated as described previously (Chen et al, ). Hydrazide linkages for conjugating reducing sugars to the poly(PFPA) were generated by incubating the poly(PFPA)‐grafted substrates in dimethylformamide (DMF) containing hydrazine monohydrate and trimethylamine (Chen et al, ). We controlled the density of hydrazide linkers by replacing various percentages of hydrazine with ethanolamine hydrochloride.…”

“…In the first model, we utilized sialylated glycoproteins bound to an inert surface in order to extend the findings of Roberts et al () to include the analysis of gliding motility. The second model employed a protocol for surface‐grafted glycopolymer construction (Chen et al, ) that allowed us to control precisely the oligosaccharide chemical presentation and surface grafting density. In both models, the surface density of sialylated residues influenced M. pneumoniae gliding frequency but not gliding speed.…”