binding constants of levans and D-fructo-oligosaccharides to BALB/c and NZB D-fructan-specific, myeloma proteins, determined by affinity electrophoresis

Abstract

The association constants for the interaction of BALB/c (UPC 10, Y5476, W3082, and UPC 61) and NZB (PC 3660) myeloma anti-D-fructans in pure form, or in ascitic fluids, with high-molecular-weight levans (Ka) and with such low-molecular-weight compounds as rye-grass levan, inulin, sucrose, and D-fructo-oligosaccharides (Kia) were determined by affinity electrophoresis, measuring the extent of retardation of the D-fructan-specific band by levan and its restoration by the low-molecular-weight compounds and oligosaccharide haptens. With different levans,Ka values ranged from 1.14 X 10(5) TO 1.52 X 10(6) ML/g for PC 3660, 1.35 X 10(5) TO 6.45 X 10(5) mL/g for UPC 10, 1.0 X 10(4) to 7.9 X 10(4) mL/g for Y5476, 7.63 X 10(3) to 6.38 X 10(4) mL/g for W 3082, and 3.35 X 10(3) to 1.54 X 10(4) mL/g for UPC 61. The retarded, D-fructan-specific bands of W3082 and UPC 61 were restored by inulin, having beta-D-(2 leads to)-linkages, and rye-grass levan, having beta-D-(2 leads to 6)-linkages, and those of PC 3660, UPC 10, and Y5476 by rye-grass levan. The Kia values of inulin with W3082 and UPC 61 were 10 times those of rye-grass levan. The Kia values of inulin (3.65 X 10(5) M-1 for W3082, and 4.44 X 10(5) M-1 for UPC 61) were very similar to those of [beta-D-Fruf-(2 leads to 1)]2-beta-D-Fruf-(2 leads to 6)-D-Glc (3) (3.95 X 10(5) M-1 for W3082, and 4.5 X 10(5) M-1 for UPC 61). With sucrose and the D-fructo-oligosaccharides, the order of Kia values of W3082 and UPC 61 was 3 greater than beta-D-Fruf-(2 leads to 1)-beta-D-Fruf-(2 leads to 6)-D-Glc greater than sucrose greater than beta-D-Fruf-(2 leads to 6)-D-Glc. With rye-grass levan, Kia values were 4.25 X 10(5) M-1 for PC 3660, 2.7 X 10(5) M-1 for UPC 10, and 8.77 X 10(4) M-1 for Y5476. These results confirm earlier findings that W3082 and UPC 61 have dual specificity for beta-(2 leads to 1) and beta-(2 leads to 6) D-fructofuranosyl linkages, that PC 3660, Y5476, and UPC 1U have specificity for beta-(2 leads to 6) D-fructofuranosyl linkages, and that the combining sites of W3082 and UPC 61 are most complementary to the tetrasaccharide 3. That W3082 and Y5476 share the same, cross-reacting idiotype, although their combining sites differ in specificity, provides further evidence that these two properties do not run parallel.