Aberrant glycosylation of cell membrane and secreted glycoproteins is a hallmark of various disease states, including cancer. The natural lectins currently used in the recognition of these glycoproteins are costly, difficult to produce, and unstable toward rigorous use. Herein we describe the design and synthesis of several boronic acid functionalized peptide-based synthetic lectin (SL) libraries, as well as the optimized methodology for obtaining peptide sequences of these SLs. SL libraries were subsequently used to identify SLs with as high as 5-fold selectivity for various glycoproteins. SLs will inevitably find a role in cancer diagnostics, given that they do not suffer from the drawbacks of natural lectins and that the combinatorial nature of these libraries allows for the identification of an SL for nearly any glycosylated biomolecule.

The binding of 125I-labeled nerve growth factor (NGF) to human melanoma cell (A875) membranes, detergent-soluble membrane extracts, and membrane extracts reconstituted into phospholipid vesicles was significantly increased when binding was carried out in the presence of wheat germ agglutinin (WGA). In the absence of WGA, all 125I-NGF binding was rapidly eliminated by trypsin treatment or rapidly dissociated in the presence of a high concentration of unlabeled NGF. However, in the presence of WGA, up to 75% of 125I-NGF bound was resistant to trypsin digestion and was only slowly dissociated by a high concentration of unlabeled NGF. The effects of WGA can be blocked or reversed by N-acetylglucosamine. Both WGA and NGF rapidly associate with soluble extracts and reconstituted vesicles and, at the concentrations used here, reach binding equilibrium within 2 min. The conversion to slowly dissociating, trypsin-resistant binding, however, was not complete for at least 10 min. Both WGA and NGF are required for maximum accumulation of trypsin-resistant, slowly dissociating binding. The order of addition of NGF and WGA has no effect on the rate of conversion of NGF-receptor, and the conversion occurs after both NGF and WGA are present. The amount of conversion is dependent on the incubation temperature, and significantly greater conversion occurs at 37 than at 0 degrees C. The generation of the trypsin-resistant, slowly dissociating state of NGF-receptor is consistent with a time- and temperature-dependent conformational change in NGF-receptor which occurs after interaction of both NGF and WGA with the receptor or closely associated structures.

Cells express many proteins that bind to laminin, the major adhesive component of basement membranes. Some of these, specifically integrins, function as transmembrane receptors that 'signal' the presence of laminin on the cell surface to the cytoplasm. Lectins constitute a second class of laminin binding proteins that may augment integrin function by interacting with laminin carbohydrate. Caution must be used in ascribing functions to other laminin binding proteins, especially cytosolic proteins.

N-Glycans of Entamoeba histolytica, the protist that causes amebic dysentery and liver abscess, are of great interest for multiple reasons. E. histolytica makes an unusual truncated N-glycan precursor (Man(5)GlcNAc(2)), has few nucleotide sugar transporters, and has a surface that is capped by the lectin concanavalin A. Here, biochemical and mass spectrometric methods were used to examine N-glycan biosynthesis and the final N-glycans of E. histolytica with the following conclusions. Unprocessed Man(5)GlcNAc(2), which is the most abundant E. histolytica N-glycan, is aggregated into caps on the surface of E. histolytica by the N-glycan-specific, anti-retroviral lectin cyanovirin-N. Glc(1)Man(5)GlcNAc(2), which is made by a UDP-Glc: glycoprotein glucosyltransferase that is part of a conserved N-glycan-dependent endoplasmic reticulum quality control system for protein folding, is also present in mature N-glycans. A swainsonine-sensitive alpha-mannosidase trims some N-glycans to biantennary Man(3)GlcNAc(2). Complex N-glycans of E. histolytica are made by the addition of alpha1,2-linked Gal to both arms of small oligomannose glycans, and Gal residues are capped by one or more Glc. In summary, E. histolytica N-glycans include unprocessed Man(5)GlcNAc(2), which is a target for cyanovirin-N, as well as unique, complex N-glycans containing Gal and Glc.