MOLECULAR DYNAMICS-DERIVED CONFORMATION AND INTRAMOLECULAR INTERACTION ANALYSIS OF THE N-ACETYL-9-O-ACETYLNEURAMINIC ACID-CONTAINING GANGLIOSIDE G(D1A) AND NMR-BASED ANALYSIS OF ITS BINDING TO A HUMAN POLYCLONAL IMMUNOGLOBULIN-G FRACTION WITH SELECTIVITY FOR O-ACETYLATED SIALIC ACIDS

The influence of 9-O-acetylation of G(D1a), yielding G(D1a) ((e)Neu5,9 Ac(2)) with a 9-O-acetylated sialic acid moiety linked to the outer ga lactose residue, on the spatial extension and mobility of the carbohyd rate chain and on recognition by a natural human antibody is analysed, To study a potential impact of the O-acetyl group on the overall conf ormation of the carbohydrate chain, molecular dynamics (MD) simulation s of oligosaccharide chain fragments of increasing length starting fro m the non-reducing end have been carried out for the first time in thi s study. They revealed a considerable loss in chain flexibility after addition of the internal N-acetylneuraminic acid onto the chain, Besid es MD calculations with different dielectric constants, the conformati onal behaviour of the complete oligosaccharide chain of the 9-O-acetyl ated G(D1a) ganglioside was simulated in the solvents water and dimeth yl sulfoxide, These solvents were also used in NMR measurements, The r esults of this study indicate that 9-O-acetylation at the terminal sia lic acid does not influence the overall conformation of the gangliosid e, An extended interaction analysis of energetically minimized conform ations of G(D1a) ((e)Neu5,9Ac(2)) and G(D1a), obtained during molecula r dynamics simulations, allowed assessment of the influence of the dif ferent parts of the saccharide chains on spatial flexibility, Notewort hy energetic interactions, most interestingly between the 9-O-acetyl g roup and the pyranose ring of N-acetylgalactosamine, were ascertained by the calculations. However, the strength of this interaction does no t force the ganglioside into a conformation, where the 9-O-acetyl grou p is no longer accessible, Binding of G(D1a) ((e)Neu5,9Ac(2)) to prote ins, which are specific for 9-O-acetylated sialic acids, should thus a t least partially be mediated by the presence of this group, To experi mentally prove this assumption, a NMR study of 9-O-acetylated G(D1a) i n the presence of an affinity-purified polyclonal IgG fraction from hu man serum with preferential binding to 9-O-acetylated sialic acid was performed, The almost complete disappearance of the intensity of the 9 -O-acetyl methyl signal of the G(D1a) ((e)Neu5,9Ac(2)) clearly indicat es that the assumed interaction of the 9-O-acetyl group with the human protein takes place.