A protein superfamily with a "Domain of Unknown Function,", DUF3349 (PF11829), is present predominately in Mycobacterium and Rhodococcus bacterial species suggesting that these proteins may have a biological function unique to these bacteria. We previously reported the inaugural structure of a DUF3349 superfamily member, Mycobacterium tuberculosis Rv0543c. Here, we report the structures determined for three additional DUF3349 proteins: Mycobacterium smegmatis MSMEG_1063 and MSMEG_1066 and Mycobacterium abscessus MAB_3403c. Like Rv0543c, the NMR solution structure of MSMEG_1063 revealed a monomeric five α-helix bundle with a similar overall topology. Conversely, the crystal structure of MSMEG_1066 revealed a five α-helix protein with a strikingly different topology and a tetrameric quaternary structure that was confirmed by size exclusion chromatography. The NMR solution structure of a fourth member of the DUF3349 superfamily, MAB_3403c, with 18 residues missing at the N-terminus, revealed a monomeric α-helical protein with a folding topology similar to the three C-terminal helices in the protomer of the MSMEG_1066 tetramer. These structures, together with a GREMLIN-based bioinformatics analysis of the DUF3349 primary amino acid sequences, suggest two subfamilies within the DUF3349 family. The division of the DUF3349 into two distinct subfamilies would have been lost if structure solution had stopped with the first structure in the DUF3349 family, highlighting the insights generated by solving multiple structures within a protein superfamily. Future studies will determine if the structural diversity at the tertiary and quaternary levels in the DUF3349 protein superfamily have functional roles in Mycobacteria and Rhodococcus species with potential implications for structure-based drug discovery.

中文翻译：

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分枝杆菌DUF3349超家族中的结构多样性。

具有“未知功能域”的蛋白质超家族DUF3349（PF11829）主要存在于分枝杆菌属和红球菌属细菌物种中，表明这些蛋白质可能具有这些细菌特有的生物学功能。我们之前报道了DUF3349超家族成员结核分枝杆菌Rv0543c的就职结构。在这里，我们报告确定了三个附加DUF3349蛋白的结构：耻垢分枝杆菌MSMEG_1063和MSMEG_1066和脓肿分枝杆菌MAB_3403c。与Rv0543c一样，MSMEG_1063的NMR溶液结构显示出具有相似总体拓扑结构的单体5α-螺旋束。反过来，MSMEG_1066的晶体结构揭示了五个具有显着不同拓扑结构的α-螺旋蛋白和一个四聚体四级结构，该结构已通过尺寸排阻色谱法确认。DUF3349超家族的第四个成员MAB_3403c的NMR溶液结构在N末端缺失18个残基，揭示出一种单体α螺旋蛋白，其折叠拓扑类似于MSMEG_1066起始子中的三个C端螺旋。四聚体。这些结构，再加上基于GREMLIN的DUF3349主要氨基酸序列的生物信息学分析，提示了DUF3349家族中的两个亚家族。如果DUF3349系列中的第一个结构停止了结构求解，则DUF3349分为两个不同的子族的划分将会丢失。强调通过解决蛋白质超家族中的多个结构而产生的见解。未来的研究将确定DUF3349蛋白超家族中第三级和第四级的结构多样性是否在分枝杆菌属和红球菌属中具有功能性作用，对基于结构的药物发现具有潜在的影响。