Effect of Globotriaosyl Ceramide Fatty Acid α-Hydroxylation on the Binding by Verotoxin 1 and Verotoxin 2.

Variation in the lipid moiety of the verotoxin (VT) receptor glycosphingolipid, globotriaosyl ceramide (Gb[sub 3]) can modulate toxin binding. The binding of VT1 and VT2 to C18 and C22 αhydroxy and nonhydroxy fatty acid isoforms of Gb[sub 3] were compared using a receptor ELISA and a [sup 125]I-labeled toxin/glycolipid microtitre plate direct binding assay. Increased binding to the hydroxylated species, particularly C22OH, was observed for both toxins. Increased RELISA binding at low glycolipid concentrations only, suggested the binding affinity is increased following Gb[sub 3] fatty acid hydroxylation. Nonlinear regression analysis of direct binding assay to these Gb[sub 3] isoforms confirmed the increased affinity of both toxins for the C22 hydroxylated Gb[sub 3]. The capacity was also significantly increased. The increased binding of VTs for hydroxylated fatty acid Gb[sub 3] isoforms may be a factor in the selective renal pathology which can follow systemic verotoxemia, particularly in the mouse model. The more pronounced effect at lower glycolipid concentrations prompted investigation of VT1 binding affinity at different Gb[sub 3] concentrations. Unexpectedly, the VT1 Kd for Gb[sub 3] was found to decrease as an inverse function of the Gb[sub 3] concentration. This shows that glycolipids have "nonclassical" receptor properties. [ABSTRACT FROM AUTHOR]