Yaniv Oren, Frolow Felix, Levy-Assraf Maly, Lamed Raphael, Bayer Edward A
Department of Molecular Microbiology and Biotechnology, The Daniella Rich Institute for Structural Biology, Tel Aviv University, Ramat Aviv, Israel.
Methods Enzymol. 2012;510:247-59. doi: 10.1016/B978-0-12-415931-0.00013-6.
Family 3 carbohydrate-binding modules (CBM3s) are among the most distinctive, diverse, and robust. CBM3s, which are numerous components of both free cellulases and cellulosomes, bind tightly to crystalline cellulose, and thus play a key role in cellulose degradation through their substrate targeting capacity. In addition to the accepted cellulose binding surface of the CBM3 molecule, a second type of conserved face (the "shallow groove") is retained on the opposite side of the molecule in all CBM3 subfamilies, irrespective of the loss or modification of the cellulose-binding function. The exact function of this highly conserved shallow groove is currently unknown. The cellulosomal system contains many linker segments that interconnect the various modules in long polypeptides chains. These linkers are varied in length (5-700 residues). The long linkers are commonly composed of repeated sequences that are often rich in Ser, Pro, and Thr residues. The exact function of the linker segments in the cellulosomal system is currently unknown, although they likely play several roles. In this chapter, we document the binding interaction between the conserved shallow-groove region of the CBM3s with selected cellulosomal linker segments, which may thus induce conformational changes in the quaternary structure of the cellulosome. These conformational changes would presumably promote changes in the overall arrangement of the cellulosomal enzymes, which would in turn serve to enhance cellulosome efficiency and degradation of recalcitrant polysaccharide substrates. Here, we describe two different methods for determining the interactions between a model CBM3 and cellulosomal linker peptides.
家族3碳水化合物结合模块（CBM3s）是最具特色、最多样化且最稳定的模块之一。CBM3s是游离纤维素酶和纤维小体的众多组成部分，能紧密结合结晶纤维素，因此通过其底物靶向能力在纤维素降解中发挥关键作用。除了公认的CBM3分子纤维素结合表面外，在所有CBM3亚家族中，分子的另一侧还保留着第二种保守面（“浅沟”），无论纤维素结合功能是否丧失或改变。这种高度保守的浅沟的确切功能目前尚不清楚。纤维小体系统包含许多连接片段，这些片段在长多肽链中连接各个模块。这些连接子长度各异（5 - 700个残基）。长连接子通常由富含丝氨酸、脯氨酸和苏氨酸残基的重复序列组成。纤维小体系统中连接片段的确切功能目前尚不清楚，尽管它们可能发挥多种作用。在本章中，我们记录了CBM3s保守浅沟区域与选定的纤维小体连接片段之间的结合相互作用，这可能会诱导纤维小体四级结构的构象变化。这些构象变化可能会促进纤维小体酶整体排列的改变，进而提高纤维小体效率和对难降解多糖底物的降解能力。在此，我们描述了两种不同的方法来确定模型CBM3与纤维小体连接肽之间的相互作用。
