The binding of the catalytic domain of Thermobifida fusca endoglucanase (EC 3.2.1.4) CDCel5A and exoglucanases (EC 3.2.1.91) CDCel6B and CDCel48A to BMOC was studied. At 5degreesC, the binding of these CDs to BMCC is rapid with maximum binding occurring just after CD loading. An observed desorption of the bound CDs with time was attributed to the loss of binding sites due to hydrolysis of the easily hydrolysable BMOC. This conclusion was supported by prehydrolysis experiments where the easily hydrolysable BMCC fraction was removed, and binding to the recalcitrant fraction was observed. More of the CDs were bound to the recalcitrant fraction. This was especially true for the exoglucanases, CDCel6B and CDCel48A. where 81-76% of the total CD binding was to the recalcitrant BMCC fraction. The binding to the easily hydrolysable fraction for the endo CDCel5A was 1.5-4 times higher than that of the exo CDCel6B and CDCel48A at the same enzyme concentration. Although CDCel5A saturated the easily hydrolysable fraction of BMCC first, the ratio of the bound to the recalcitrant substrate was constant for CDCel6B and CDCel48A as 81 and 76%, respectively. As the reaction temperature was increased to 50degreesC, CDCel5A and CDCel48A exhibited a rapid increase in the rate and extent of binding, while CDCel6B exhibited a decrease in the extent of binding.