On the structural significance of the linkage region constituents of N-glycoproteins: an X-ray crystallographic investigation using models and analogs

Abstract

The linkage region constituents, namely, 2-acetamido-2-deoxy-β- d-glucopyranose and asparagine are conserved in the N-glycoproteins of all the eukaryotes. The present work is aimed at understanding the reasons for the occurrence of GlcNAc and Asn as the linkage region constituents. A total of six sugar amides have been designed as models and analogs of the linkage region and their crystal structures have been solved. This is the first report on the X-ray crystallographic investigation of the effect of systematic changes in the linkage sugar as well as its aglycon moiety on the N-glycosidic torsion, ψ N (O5–C1–N1–C1 ′). This also forms the first report on the crystal structure of a model of l-RhaβAsn, a variant linkage found in the surface layer glycoprotein of Bacillus stearothermophillus. Among the models and analogs examined, the acetamido derivatives of Man and Xyl, the linkage sugars of O-glycoproteins, show a ψ N value of −114.5° and −121.2°, respectively, deviating maximum from the value of −89.8° reported for the model compound GlcNAcβNHAc. The l-Rha and Gal derivatives also show noticeable deviations. The ψ N values, −89.5° and −91.0°, of the propionamide derivatives of Glc and GlcNAc (analogs of GlcβGln and GlcNAcβGln, respectively) agree well with those (−93.8° and −89.8°) reported for their corresponding acetamide derivatives suggesting Gln could serve as well as Asn as the linkage region amino acid. However, the rotational freedom about the additional C–C bond would lead to altered rigidity of the linkage region. An analysis of packing reveals that the molecular assembly of these compounds is driven by different infinite and finite chains of hydrogen bonds. The double pillaring of hydrogen bonds involving the amide groups at C1 and C2 is seen as a unique packing feature characteristic of β-1- N-acyl derivatives of GlcNAc. Based on the findings of the present study, it is speculated that the linkage region constituents of the eukaryotic N-glycoproteins appear to fulfill three essential structural requirements: rigidity, planarity, and linearity and these are met by the trisaccharide core and Asn at the linkage region.