The O-
glycans in
Jacalin-binding
immunoglobulin A1 (
IgA1) were released by gas-phase hydrazinolysis and were fractionated by gel filtration and reversed-phase HPLC. Four peaks (P1 to P4) were obtained. There was a significant shift from Peak 2 (monosialylated Gal beta 1-3GalNAc) to Peak 4 (asialo-Gal beta 1-3GalNAc) in the
IgA-nephropathy group compared with the negative control group (P < 0.05). One of the functions of the
carbohydrate side chains is to stabilize the three-dimensional structure of the
glycopeptides. In order to evaluate the stability of the
Jacalin-binding
IgA1 molecule, the increase in turbidity as a consequence of the increase of the aggregated
IgA1 level under the condition of high temperature (63 degrees C) was observed. The increase in the turbidity was significantly higher in the
IgA nephropathy group compared with the negative control group (21.7 vs. 5.9% at 150 min, P < 0.02). From a sample of
IgA1 solution that had originated from a pooled normal serum, heat-tolerant (nonaggregated) and intolerant (aggregated)
IgA1 molecules were separated by gel filtration. The heat-intolerant
IgA1 had lower amounts of
sialic acid (27.4 micrograms/mg
IgA1) than the tolerant
IgA1 (37.6 micrograms/mg
IgA1). Further, the analysis of the O-
glycans released from another
IgA1 sample by the hydrazinolysis revealed that the ratio of the asialo-Gal beta 1-3GalNAc/total Gal beta 1-3GalNAc in the heat-in-tolerant
IgA1 (27.2%) was increased compared with that in the heat-tolerant
IgA1 (18.2%). From these results, it was suggested that unusual glycosylation on the hinge region of
Jacalin-binding
IgA1 induced an insufficient conformational stiffness to the hinge
peptide, resulting in the aggregation of the
IgA1 molecule in
IgA nephropathy.