Characterization of the microheterogeneity in glycoproteins by 500-MHz 1H-NMR spectroscopy of glycopeptide preparations. Application to a monofucosylated tetra-antennary glycopeptide fraction from human plasma alpha 1-acid glycoprotein.

Five hundred-MHz 1H-NMR spectroscopy was employed to study a monofucosylated tetra-antennary glycopeptide fraction which was derived from human plasma alpha 1-acid glycoprotein. This fraction was earlier judged to be homogeneous by 360-MHz 1H-NMR spectroscopic analysis (Fournet, B., Montreuil, J., Strecker, G., Dorland, L., Haverkamp, J., Vliegenthart, J. F. G., Binette, J. P., and Schmid, K. (1978) Biochemistry 17, 5206-5214). The combination of the improved resolving power and the enhanced sensitivity of the 500-MHz 1H-NMR spectrometer afforded the elucidation of a new type of microheterogeneity with regard to the position of attachment of Fuc. Three isomeric compounds were identified. The major form contains Fuc alpha-(1 leads to 3) linked to GlcNac 7 of the tetra-antennary structure, as shown earlier. The two minor compounds, representing new structures, possess Fuc attached in alpha-(1 leads to 3) linkage to GlcNAc 7' or 5'. It is thus noteworthy that this spectral technique allows elucidation of structures of very closely related carbohydrate chains in a glycopeptide mixture.