The the family 8 of glycoside hydrolases includes enzymes with more diversified substrate specificities than families GH46, GH75 or GH80. Besides chitosanases, GH8 includes cellulases, licheninases, endo-1,4-β-xylanases and reducing end xylose-releasing exo-oligoxylanases.
So far, chitosanase activity has been experimentally demonstrated for more than twenty enzymes of this family.
Chitosanase from Bacillus sp. No.7-M (27) and from Bacillus sp. K17 (115) were shown to cleave essentially only the GlcN-GlcN linkage and were classified in the Group II according to their cleavage specificity.
The resolution of the 3D structure and studies on catalytic residues of chitosanase ChoK from Bacillus sp. K17 revealed several interesting points (115). Structural comparison between ChoK and CelA (cellulase of Clostridium thermocellum NCIB 10682/JW20, another GH8 enzyme) and structure-guided alignments with sequences of other GH8 members suggested that this family can be divided into three subfamilies (named GH8a, GH8b and GH8c), depending on the position of the general base residue. GH8a members have an aspartate as the general base. In GH8b enzymes, the Asp residue is inactivated by a substitution into an asparagine, and the general base is a glutamate residue. The subfamily GH8c is similar to the subfamily 8b: the general base aspartate of the subfamily 8a is also inactivated via a substitution by an asparagine. Although no work has clearly identified the general base in this subfamily, a conserved carboxylic residue is a candidate for this function. Below we show the alignment of 11 enzymes belonging to family GH8.
It appears that there is a correlation between the subdivision into GH8a, GH8b and GH8c and the substrate specificity of the respective enzymes. GH8a includes cellulases,
xylanases and several endoglucanases. GH8b includes chitosanases, lichenases and some other endoglucanases, while GH8c groups the remaining specificities
Other extensively studied GH8 chitosanases are produced by Bacillus circulans WL-12 (8) and Paenibacillus fukuinensis strain D2 (63). Zitouni et al. also identified a highly thermostable GH8 chitosanase produced by Paenibacillus sp. 1794 isolated from compost (117).