Extra tyrosine in the carbohydrate-binding module of <i>Irpex lacteus</i> Xyn10B enhances its cellulose-binding ability

The xylanase (Xyn10B) that strongly adsorbs on microcrystalline cellulose was isolated from Driselase. The Xyn10B contains a Carbohydrate-binding module family 1 (CBM1) (<i>Irp</i>CBM_Xyn10B) at N-terminus. The canonical essential aromatic residues required for cellulose binding were conserved in <i>Irp</i>CBM_Xyn10B; however, its adsorption ability was markedly higher than that typically observed for the CBM1 of an endoglucanase from <i>Trametes hirsuta</i> (<i>Th</i>CBM_EG1). An analysis of the CBM-GFP fusion proteins revealed that the binding capacity to cellulose (7.8 μmol/g) and distribution coefficient (2.0 L/μmol) of <i>Irp</i>CBM_Xyn10B-GFP were twofold higher than those of <i>Th</i>CBM_EG1-GFP (3.4 μmol/g and 1.2 L/μmol, respectively), used as a reference structure. Besides the canonical aromatic residues (W24-Y50-Y51) of typical CBM1-containing proteins, <i>Irp</i>CBM_Xyn10B had an additional aromatic residue (Y52). The mutation of Y52 to Ser (<i>Irp</i>CBM_Y52S-GFP) reduced these adsorption parameters to 4.4 μmol/g and 1.5 L/μmol, which were similar to those of <i>Th</i>CBM_EG1-GFP. These results indicate that Y52 plays a crucial role in strong cellulose binding.从Driselase中分离得到了一种可强效吸附于微晶纤维素的木聚糖酶（Xyn10B）。该Xyn10B的N末端带有碳水化合物结合模块家族1（Carbohydrate-binding module family 1，CBM1），记为*Irp*CBM_Xyn10B。纤维素结合所必需的典型芳香族残基在*Irp*CBM_Xyn10B中均保持保守，但该模块的吸附能力显著高于毛栓菌（*Trametes hirsuta*）内切葡聚糖酶的CBM1（记为*Th*CBM_EG1）。对CBM-绿色荧光蛋白（Green Fluorescent Protein，GFP）融合蛋白的分析结果显示，*Irp*CBM_Xyn10B-GFP的纤维素结合容量（7.8 μmol/g）与分布系数（2.0 L/μmol）分别为参照组*Th*CBM_EG1-GFP（对应值分别为3.4 μmol/g和1.2 L/μmol）的2倍。除典型含CBM1蛋白的保守芳香族残基（W24-Y50-Y51）外，*Irp*CBM_Xyn10B还额外带有一个芳香族残基Y52。将Y52突变为丝氨酸后得到突变体*Irp*CBM_Y52S-GFP，其吸附参数降至4.4 μmol/g和1.5 L/μmol，与*Th*CBM_EG1-GFP的对应参数相近。上述结果表明，Y52在强效纤维素结合过程中发挥关键作用。