Functional Analysis of Molybdopterin Biosynthesis in Mycobacteria Identifies a Fused Molybdopterin Synthase in Mycobacterium tuberculosis
Functional Analysis of Molybdopterin Biosynthesis in Mycobacteria Identifies a Fused Molybdopterin Synthase in Mycobacterium tuberculosis
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DOI：
10.1128/jb.00774-10
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发表时间：
2011-01-01
影响因子：
3.2
通讯作者：
Mizrahi, Valerie
中科院分区：
文献类型：
--
作者：
Williams, Monique J.;Kana, Bavesh D.;Mizrahi, Valerie
大多数分枝杆菌物种都具有完整的钼辅因子(MoCo)生物合成基因。然而，结核分枝杆菌复合体成员的一个显著特征是他们拥有与MoCo生物合成途径的前两步相关的多个同源物。缺乏moaA1-moaD1基因簇的结核分枝杆菌突变体和其中moaD2也缺失的衍生物在以硝酸盐为唯一氮源的培养基中生长显著受损，表明MoCo的可用性降低，以支持依赖MoCo的硝酸还原酶NarGHI的同化功能。然而，双突变株显示了呼吸硝酸还原酶的残留活性，这表明它保持了生产MoCo的能力。通过在耻垢分枝杆菌的突变株中表达这些基因，进一步分析了结核分枝杆菌的moad和moaE同源物，这些突变株缺乏唯一的MoaD2和moaE2合成酶编码基因中的一个或两个，并且不能生长在硝酸盐上，可能是由于失去了依赖MoCo的硝酸盐同化活性。在耻垢分枝杆菌moaD2突变体中表达moaD2和在耻垢分枝杆菌moaE2突变体中表达moaE1或moaE2恢复了硝酸盐同化，证实了这些基因在MPT合成中的功能。在缺乏moaD2和/或moaE2的耻垢分枝杆菌中，moaX的表达也恢复了MoCo的生物合成，从而证明MoaX是一种融合的MPT合成酶。通过在MoCo生物合成中涉及多个合成酶编码同源物，这些结果表明它们在结核分枝杆菌中的扩张可能服务于重要的细胞功能。
Most mycobacterial species possess a full complement of genes for the biosynthesis of molybdenum cofactor (MoCo). However, a distinguishing feature of members of the Mycobacterium tuberculosis complex is their possession of multiple homologs associated with the first two steps of the MoCo biosynthetic pathway. A mutant of M. tuberculosis lacking the moaA1-moaD1 gene cluster and a derivative in which moaD2 was also deleted were significantly impaired for growth in media containing nitrate as a sole nitrogen source, indicating a reduced availability of MoCo to support the assimilatory function of the MoCo-dependent nitrate reductase, NarGHI. However, the double mutant displayed residual respiratory nitrate reductase activity, suggesting that it retains the capacity to produce MoCo. The M. tuberculosis moaD and moaE homologs were further analyzed by expressing these genes in mutant strains of M. smegmatis that lacked one or both of the sole molybdopterin (MPT) synthase-encoding genes, moaD2 and moaE2, and were unable to grow on nitrate, presumably as a result of the loss of MoCo-dependent nitrate assimilatory activity. Expression of M. tuberculosis moaD2 in the M. smegmatis moaD2 mutant and of M. tuberculosis moaE1 or moaE2 in the M. smegmatis moaE2 mutant restored nitrate assimilation, confirming the functionality of these genes in MPT synthesis. Expression of M. tuberculosis moaX also restored MoCo biosynthesis in M. smegmatis mutants lacking moaD2, moaE2, or both, thus identifying MoaX as a fused MPT synthase. By implicating multiple synthase-encoding homologs in MoCo biosynthesis, these results suggest that important cellular functions may be served by their expansion in M. tuberculosis.