Lectin-based homogeneous enzyme-linked binding assay for estimating the type and relative amount of carbohydrate within intact glycoproteins

The feasibility of using a new lectin-based homogeneous enzyme-linked binding assay for estimating the type and relative amount of specific carbohydrate structures within intact glycoproteins is examined. Malate dehydrogenase-galactose, -mannose, and -N-acetylglucosamine conjugates are utilized in conjunction with Jacalin, concanavalin A, and wheat germ agglutinin, respectively. The catalytic activity of the glyco-enzyme conjugates is inhibited significantly (>60%) in solution in the presence of the respective lectins. The observed inhibition for each reagent set is reversed in proportion to the type and relative amount of specific carbohydrates present within test glycoproteins added to the assay mixture. Competitive binding ED50 values for a number of synthetic and native model glycoproteins correlate well with the known carbohydrate content of these species. The proposed method is much faster than previous solid-phase lectin-based enzymelinked methods used to probe carbohydrate content/structure (<15 min) and has the potential to be fully automated.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/30117/1/0000493.pd

Carbohydrate binding properties of the stinging nettle (Urtica dioica) rhizome lectin

The interaction of the stinging nettle rhizome lectin (UDA) with carbohydrates was studied by using the techniques of quantitative precipitation, hapten inhibition, equilibrium dialysis, and uv difference spectroscopy. The Carbohydrate binding site of UDA was determined to be complementary to an N,N',N"-triacetylchitotriose unit and proposed to consist of three subsites, each of which has a slightly different binding specificity. UDA also has a hydrophobic interacting region adjacent to the carbohydrate binding site. Equilibrium dialysis and uv difference spectroscopy revealed that UDA has two carbohydrate binding sites per molecule consisting of a single polypeptide chain. These binding sites either have intrinsically different affinities for ligand molecules, or they may display negative cooperativity toward ligand binding.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/26071/1/0000145.pd

N-Acetylglucosamine: Production and Applications

N-Acetylglucosamine (GlcNAc) is a monosaccharide that usually polymerizes linearly through (1,4)-β-linkages. GlcNAc is the monomeric unit of the polymer chitin, the second most abundant carbohydrate after cellulose. In addition to serving as a component of this homogeneous polysaccharide, GlcNAc is also a basic component of hyaluronic acid and keratin sulfate on the cell surface. In this review, we discuss the industrial production of GlcNAc, using chitin as a substrate, by chemical, enzymatic and biotransformation methods. Also, newly developed methods to obtain GlcNAc using glucose as a substrate in genetically modified microorganisms are introduced. Moreover, GlcNAc has generated interest not only as an underutilized resource but also as a new functional material with high potential in various fields. Here we also take a closer look at the current applications of GlcNAc, and several new and cutting edge approaches in this fascinating area are thoroughly discussed