Investigation of the effect of specific enzyme scission on MIP1α binding to HS.

³H-labeled murine fibroblast HS chains were treated with either heparinase I or heparinase III enzymes as described in “Materials and methods.” Intact (A), heparinase I-digested, (B) and heparinase III–digested (C) HS were applied to an MIP1α (BB10010) Affi-Gel column (▪) in 0.15 M NaCl. Identically prepared HS was also applied to a control column (●). Bound material was eluted with a stepwise NaCl gradient (^{. . . . . . .}). (D) Sulfated domains excised from HS by heparinase III were separated by Biogel P10 chromatography, then applied to the Affi-Gel MIP1α (BB10010) column. All the material was eluted with 0.2 M and 0.4 M NaCl steps. Most of the disaccharides and tetrasaccharides were eluted in the 0.15 M NaCl wash; hence, the bars at 0.2 M and 0.4 M are undetectable. ■ indicates hexasaccharides; ▪, octasaccharides; ▨, decasaccharides; ▩, dodecasaccharides; and ▤, tetradecasaccharides. Each graph is representative of 2 or more experiments.