The Shmunis School of Biomedicine and Cancer Research, Tel Aviv University, Ramat Aviv 69978, Israel.
The Wolfson Center for Applied and Structural Biology, Department of Biological Chemistry, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 9190401, Israel.
Acta Crystallogr F Struct Biol Commun. 2021 Apr 1;77(Pt 4):95-104. doi: 10.1107/S2053230X21002764. Epub 2021 Mar 31.
A novel member of the family 3 carbohydrate-binding modules (CBM3s) is encoded by a gene (Cthe_0271) in Clostridium thermocellum which is the most highly expressed gene in the bacterium during its growth on several types of biomass substrates. Surprisingly, CtCBM3-0271 binds to at least two different types of xylan, instead of the common binding of CBM3s to cellulosic substrates. CtCBM3-0271 was crystallized and its three-dimensional structure was solved and refined to a resolution of 1.8 Å. In order to learn more about the role of this type of CBM3, a comparative study with its orthologue from Clostridium clariflavum (encoded by the Clocl_1192 gene) was performed, and the three-dimensional structure of CcCBM3-1192 was determined to 1.6 Å resolution. Carbohydrate binding by CcCBM3-1192 was found to be similar to that by CtCBM3-0271; both exhibited binding to xylan rather than to cellulose. Comparative structural analysis of the two CBM3s provided a clear functional correlation of structure and binding, in which the two CBM3s lack the required number of binding residues in their cellulose-binding strips and thus lack cellulose-binding capabilities. This is an enigma, as CtCBM3-0271 was reported to be a highly expressed protein when the bacterium was grown on cellulose. An additional unexpected finding was that CcCBM3-1192 does not contain the calcium ion that was considered to play a structural stabilizing role in the CBM3 family. Despite the lack of calcium, the five residues that form the calcium-binding site are conserved. The absence of calcium results in conformational changes in two loops of the CcCBM3-1192 structure. In this context, superposition of the non-calcium-binding CcCBM3-1192 with CtCBM3-0271 and other calcium-binding CBM3s reveals a much broader two-loop region in the former compared with CtCBM3-0271.
一个新型的家族 3 碳水化合物结合模块（CBM3s）成员由热纤维梭菌（Clostridium thermocellum）中的一个基因（Cthe_0271）编码，该基因在细菌生长于多种生物质底物时表达量最高。令人惊讶的是，CtCBM3-0271 可以结合至少两种不同类型的木聚糖，而不是 CBM3s 常见的纤维素底物结合。CtCBM3-0271 被结晶，其三维结构被解析并精修至 1.8 Å分辨率。为了更多地了解这种 CBM3 的作用，与它的同源物（由 Clostridium clariflavum 的 Clocl_1192 基因编码）进行了比较研究，并确定了 CcCBM3-1192 的三维结构分辨率为 1.6 Å。发现 CcCBM3-1192 的碳水化合物结合与 CtCBM3-0271 的相似；两者都表现出与木聚糖结合而不是纤维素结合。对这两种 CBM3 的比较结构分析提供了结构与结合的明确功能相关性，其中这两种 CBM3 在其纤维素结合带中缺乏所需数量的结合残基，因此缺乏纤维素结合能力。这是一个谜，因为当细菌生长在纤维素上时，CtCBM3-0271 被报道为高度表达的蛋白质。一个额外的意外发现是，CcCBM3-1192 不含被认为在 CBM3 家族中起结构稳定作用的钙离子。尽管缺乏钙离子，但形成钙离子结合位点的五个残基是保守的。钙离子的缺失导致 CcCBM3-1192 结构中两个环的构象变化。在这种情况下，将非钙离子结合的 CcCBM3-1192 与 CtCBM3-0271 和其他钙离子结合的 CBM3 进行叠加，显示前者的两个环区比 CtCBM3-0271 更宽。
