Jean-Philippe Grivet

Protein—sugar interactions A nuclear magnetic resonance investigation of the binding of O‐methyl‐di‐N‐acetyl‐β‐chitobioside to wheat germ agglutinin (lectin)

Wheat germ agglutinin (WGA) is a plant lectin which can agglutinate various animal cells, particularly malignant cells and normal protease treated cells [1-3]. Agglutination is inhibited by N-acetyl-glucosamine and its/3-1--4-1inked oligomers [2--4]. Under physiological conditions, WGA is a dimer. In acidic media, it dissociates in 2 subunits mol. wt 18 000 [5,6]. The dimer binds 4 ligands containing the Naeetylglucosamine moiety with equal affinity. In other words, each protomer binds saccharides at 2 spatially different but chemically equivalent locations [5,7,8]. Lectin-sugar interactions have been studied by fluorescence [9-13], absorption [13] and circular dichroism [ 14] techniques. Although nuclear magnetic resonance (NMR) has been used extensively to study the interactions of sugars with lysozyme (see ref in [ 15 ] ) and concanavalin A [16], only one report so far has been devoted to WGA [ 17]. The binding constant ofN-acetyl glucosamine was determined [ 17] and showed that the methyl group resonance was strongly shifted upon binding. In this communication, we present results of an z H NMR investigation of the binding of 1-O-methyl-di-N-acetyl-/~-chitobioside (CBOCH3) and of 1-O-methyl-tri-N-acetyl~-chitotrioside (CTOCHs) to WGA. The use of methyloside derivatives, in effect removing the anomeric equilibrium, greatly simplifies the analysis.

The binding of monosaccharides to wheat germ agglutinin: Fluorescence and NMR investigations

The synthesis of N-acetyl- and N-trifluoroacetyl-glucosaminides was reported. The interaction of these compounds with wheat germ agglutinin, a plant lectin specific for N-acetyl-glucosamine and sialic acid, was investigated by two complementary approaches: 1H and 19F NMR, and fluorescence spectroscopy. This last technique relies on the existence of a competitive equilibrium involving the protein, the ligand and O-(methylumbelliferyl)-N-acetyl-glucosaminide, a fluorescent saccharide. The binding constants and the chemical shifts in the complex were determined and were related to the protein structure.

chapter 14 Accuracy and precision of intensity determinations in quantitative nmr

Publisher Summary This chapter discusses accuracy and precision of intensity determination in quantitative. There is no universal data handling procedure that leads to clean baselines and to accurate and precise integrals for every type of nuclear magnetic resonance (NMR) experiment. It is in fact fortunate that the application, which is the most “integral-intensive” (NOESY spectra) is also the most tolerant to errors. Semi automatic (least-squares) or automatic (linear prediction or maximum entropy) methods should be used whenever possible, as they relieve the spectroscopist of much tedious work, can be reproducible and objective, and provide estimates of probable errors. The main argument raised against them is the long computation times involved. This time should be compared to the time spent in preparing the sample, recording the data, and assigning the spectra. The ready availability of fast work stations can also make these methods accessible to a growing number of spectroscopists.

Efficiency estimation for single-coil, separate-input, double-tuned NMR probes

Abstract The signal-to-noise ratio for the NMR observation of a nucleus using a double-tuned probe depends on the power efficiency of the observe channel. In a similar manner, the minimum decoupling power is a function of the efficiency of the proton channel. Efficiencies are often determined in several steps, which involve measuring electrical characteristics of disassembled probe elements. In this work, we show how efficiencies can be simply computed from measurements of the quality factors of each port, at its resonant frequency. Thus, an index of the probe performance, especially its signal-to-noise ratio, is readily available for the complete, loaded device.

13C NMR studies of bacterial fermentations

We describe the experimental methods used and the constraints that apply in studies of anaerobic cell metabolism by 13C NMR. We review some of the results of our recent work in this area. Clostridium neopropionicum was shown to ferment ethanol into propionate by the acrylate, non-randomizing pathway. The same metabolic route accounts for 50% of the propionate formed in the complex ecosystem that inhabits the pigs large intestine. The rest is formed via the randomizing succinate pathway. Reductive, hydrogenotrophic acetogenesis was studied in several ecosystems. Although it is usually overshadowed by methanogenesis in the competition for hydrogen, it may become an efficient electron sink when methane biosynthesis is blocked by a specific inhibitor.