Department of Molecular Nutrition, CSIR-Central Food Technological Research Institute (CFTRI), Mysuru, Karnataka, 570020, India; Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, 201002, India.
Department of Molecular Nutrition, CSIR-Central Food Technological Research Institute (CFTRI), Mysuru, Karnataka, 570020, India.
Biochimie. 2020 Apr-May;171-172:136-146. doi: 10.1016/j.biochi.2020.03.002. Epub 2020 Mar 5.
The structural maintenance of chromosomes (SMC) proteins play a vital role in genome stability and chromosome organization in all domains of life. Previous reports show that smc deletion causes decondensation of chromosome and an increased frequency of anucleated cells in bacteria. However, smc deletion in both Mycobacterium smegmatis and Mycobacterium tuberculosis did not affect chromosome condensation or the frequency of anucleated cells. In an attempt to understand this difference in M. smegmatis, we investigated the function of MksB (MsMksB), an alternate SMC-like protein. Like other bacterial SMCs, MsMksB is also an elongated homodimer, in which a central hinge domain connects two globular ATPase head domains via two coiled-coil arms. We show that full-length MsMksB binds to different topological forms of DNA without any preferences. However, the hinge and headless domains prefer binding to single-stranded DNA (ssDNA) and linear double-stranded DNA (dsDNA), respectively. The binding of MsMksB to DNA was independent of ATP as its ATP hydrolysis deficient mutant was also proficient in DNA binding. Further, the cytological profiling studies revealed that only the full-length MsMksB and none of its structural domains could condense the bacterial chromosome. This observation indicates the plausibility of the concerted action of different structural domains of SMC to bind and condense the chromosome. Moreover, MsMksB exhibited DNA stimulated ATPase activity, in addition to its intrinsic ATPase activity. Taken together, we have elucidated the function of an alternate bacterial condensin protein MksB and its structural domains in DNA binding and condensation.
染色体结构维持（SMC）蛋白在所有生命领域的基因组稳定性和染色体组织中起着至关重要的作用。以前的报告表明，smc 缺失会导致染色体去凝聚，并增加细菌中非核细胞的频率。然而，在耻垢分枝杆菌和结核分枝杆菌中 smc 的缺失都不会影响染色体的凝聚或非核细胞的频率。为了试图理解耻垢分枝杆菌中这种差异，我们研究了替代 SMC 样蛋白 MksB（MsMksB）的功能。与其他细菌 SMC 一样，MsMksB 也是一个拉长的同源二聚体，其中中央铰链结构域通过两个卷曲螺旋臂将两个球形 ATP 酶头部结构域连接起来。我们表明全长 MsMksB 可以结合不同拓扑形式的 DNA，而没有任何偏好。然而，铰链和无头结构域分别优先结合单链 DNA（ssDNA）和线性双链 DNA（dsDNA）。MsMksB 与 DNA 的结合不依赖于 ATP，因为其 ATP 水解缺陷突变体也能有效地结合 DNA。此外，细胞学分析研究表明，只有全长 MsMksB，而其任何结构域都不能使细菌染色体浓缩。这一观察结果表明，SMC 的不同结构域协同作用以结合和浓缩染色体的可能性。此外，MsMksB 表现出 DNA 刺激的 ATPase 活性，除了其内在的 ATPase 活性。综上所述，我们阐明了替代细菌浓缩蛋白 MksB 及其结构域在 DNA 结合和浓缩中的功能。
