STRUCTURE OF THE ENDOGLUCANASE-I FROM FUSARIUM-OXYSPORUM - NATIVE, CELLOBIOSE, AND 3,4-EPOXYBUTYL BETA-D-CELLOBIOSIDE-INHIBITED FORMS, AT 2.3 ANGSTROM RESOLUTION

The mechanisms involved in the enzymatic degradation of cellulose are of great ecological and commercial importance. The breakdown of cellul ose by fungal species is performed by a consortium of free enzymes, kn own as cellobiohydrolases and endoglucanases, which are found in many of the 57 glycosyl hydrolase families. The structure of the endoglucan ase I (EG I), found in glycosyl hydrolase family 7, from the thermophi lic fungus Fusarium oxysporum has been solved at 2.3 Angstrom resoluti on. In addition to the native enzyme, structures have also been determ ined with both the affinity label, 3,4-epoxybutyl beta-D-cellobioside, and the reaction product cellobiose. The affinity label is covalently bound, as expected, to the catalytic nucleophile, Glu197, with clear evidence for binding of both the R and S stereoisomers. Cellobiose is found bound to the -2 and -1 subsites of the enzyme. In marked contras t to the structure of EG I with a nonhydrolyzable thiosaccharide analo g, which spanned the -2, -1, and +1 subsites and which had a skew-boat conformation for the -1 subsite sugar [Sulzenbacher, G., et al. (1996 ) Biochemistry 35, 15280-15287], the cellobiose complex shows no pyran oside ring distortion in the -1 subsite, implying that strain is induc ed primarily by the additional +1 subsite interactions and that the pr oduct is found, as expected, in its unstrained conformation.