An artificial receptor for glycoproteins

Abstract

We describe the rational design, synthesis and development of a sterilizable biomimetic ligand for the affinity purification of glycoproteins. Based on mimicking the principles of natural carbohydrate recognition, a putative library of 196 glycoprotein-binding synthetic ligands was designed and synthesized on a polymeric support. Ligand 11/11, based on a triazine scaffold and immobilized on a hydrophilic support, was identified as the "lead" ligand. The carbohydrate recognizing the potential of the "lead" ligand was revealed by reduced binding of a periodate oxidized model glycoprotein, and by "sharp" elution profiles achieved with borate buffer eluents. Specific elution and competitive binding experiments determined the monosaccharide specificity of 11/11 in the order mannoside > glucoside > galactoside. The diastereo-selective performance of ligand 11/11 was quantified and reaffirmed by analytical affinity chromatography and (1)H-NMR, in the order galactoside < glucoside < mannoside with binding affinities (K(a), M(-1)) in the 63-214 and 20-83 M(-1) range, respectively. Partition coefficient analysis revealed binding constants towards glycoproteins in the 10(4) M(-1) range, that compared favourably with the affinities of carbohydrate binding lectins for glycoproteins, such as concanavalin A. Molecular modelling studies of ligand 11/11 revealed the formation of a pre-organized apolar "tweezer-like" cavity, containing complementary nitrogenous hydrogen bond donor and acceptor groups that formed selective interactions with the equatorial 3- and 4-hydroxyl groups of saccharides.

The Improvement of LC-MS/MS Proteomic Detection with Biomimetic Affinity Fractionation

“…As a new affinity fraction, biomimetic chromatography uses synthetic affinity ligands for matrix, which could mimic the properties of natural ligands and circumvent these drawbacks of natural ligands by imparting resistance to chemical and biochemical degradation, displaying ease and low cost of production, and withstanding harsh sterilization without loss of performance (Lowe, 2001). It has been verified that many specific synthetic affinity ligands could be designed to purify different single proteins in our lab and other labs (Li et al, 1998;Teng et al, 1999;Gupta and Lowe, 2004;Melissis et al, 2007;Xin et al, 2007;Dong et al, 2008a). Our lab has constructed affinity ligand library (thousands of ligands), and different ligands showed different absorbance effect to proteins such as purification, depletion of high abundance proteins, and enrichment of low abundance proteins (Dong et al, 2008b), which gives a imaginable application prospect in proteomics.…”

Comparison of Fluorescence Labelling Techniques for the Selection of Affinity Ligands from Solid-Phase Combinatorial Libraries

“…They tend to combine molecular recognition features with high resistance to chemical and biological degradation, high scalability, as well as low costs and low toxicity (1,3,4). A number of synthetic affinity ligands have been developed including the biomimetic or de novo designed ligands targeted at specific proteins and based on the triazine scaffold (5)(6)(7)(8)(9)(10)(11)(12). Recently, affinity ligands for immunoglobulins and their fragments based on the Ugi reaction scaffold have also been developed (13).…”

A novel fractionation method prior to MS-based proteomics analysis using cascade biomimetic affinity chromatography

“…These ligands circumvent the drawbacks of natural ligands by imparting resistance to chemical and biochemical degradation, displaying ease and low cost of production, and withstanding harsh sterilization without loss of performance. Artificial affinity ligands that mimic the properties of natural ligands have been designed and used to purify single protein [15][16][17][18][19][20]. Our lab has constructed a large affinity ligand library composed of thousands of ligands with different protein absorbance effects.…”