Jin-Tian Xu, Ji-Fang Yu, Tao Cheng, Ao Feng, Ping Yang, Jing Gu, Hong-Jun Yu, Jiao-Yu Deng
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The results of AlphaFold2 prediction indicated that the T120P or M172V mutation might affect the enzymatic activity of Rv2172c by influencing nicotinamide adenine dinucleotide (NADH) binding, and this was verified by subsequent biochemical analysis, demonstrating the role of residues Thr120 and Met172 on NADH binding and enzymatic activity of Rv2172c. In addition, the effect of <i>rv2172c</i> T120P or M172V mutation on methionine production and PAS resistance was determined in <i>M. tuberculosis</i>. The results showed that both T120P and M172V mutations caused increased intracellular methionine concentrations and high level PAS resistance. 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引用次数: 0
摘要
摘要尽管几十年来对氨基水杨酸(PAS)一直被用于治疗结核病,但结核分枝杆菌(M. tuberculosis)临床分离株对这种药物的耐药机制尚未得到深入研究。此前,我们发现 Rv2172c 的亚甲基四氢叶酸还原酶(MTHFR)活性降低会导致结核分枝杆菌对抗叶酸类药物的敏感性增加。在这项研究中,我们收集了 173 株对 PAS 耐药和 803 株对 PAS 敏感的临床分离株的基因组测序数据,并分析了这 976 株分离株中的 rv2172c 突变。结果表明,在一定比例(6.36%)的 PAS 耐药分离株中可发现 rv2172c 上的两个突变(T120P 和 M172 V)。AlphaFold2 预测结果表明,T120P 或 M172 V 突变可能通过影响烟酰胺腺嘌呤二核苷酸(NADH)的结合而影响 Rv2172c 的酶活性,这一点在随后的生化分析中得到了验证,证明了 Thr120 和 Met172 残基对 Rv2172c 的 NADH 结合和酶活性的作用。此外,还测定了 rv2172c T120P 或 M172 V 突变对结核杆菌蛋氨酸产生和 PAS 抗性的影响。结果表明,T120P 和 M172 V 突变都会导致细胞内蛋氨酸浓度增加,并产生高水平的 PAS 抗性。总之,我们在结核杆菌临床分离株中发现了新的分子标记和PAS耐药性的新机制,拓宽了对结核杆菌中Rv2172c的NADH依赖性MTHFR催化机制的认识,这将有助于PAS耐药性的分子诊断和以Rv2172c为靶点的新药开发。
The T120P or M172V mutation on rv2172c confers high level para-aminosalicylic acid resistance in Mycobacterium tuberculosis.
Although para-aminosalicylic acid (PAS) has been used to treat tuberculosis for decades, mechanisms of resistance to this drug in Mycobacterium tuberculosis (M. tuberculosis) clinical isolates have not been thoroughly investigated. Previously, we found that decreased methylenetetrahydrofolate reductase (MTHFR) activity of Rv2172c led to increased sensitivity to antifolates in M. tuberculosis. In this study, we collected the genome-sequencing data of 173 PAS-resistant and 803 PAS-sensitive clinical isolates and analyzed rv2172c mutations in those 976 isolates. The results showed that two mutations (T120P and M172V) on rv2172c could be identified in a certain proportion (6.36%) of PAS-resistant isolates. The results of AlphaFold2 prediction indicated that the T120P or M172V mutation might affect the enzymatic activity of Rv2172c by influencing nicotinamide adenine dinucleotide (NADH) binding, and this was verified by subsequent biochemical analysis, demonstrating the role of residues Thr120 and Met172 on NADH binding and enzymatic activity of Rv2172c. In addition, the effect of rv2172c T120P or M172V mutation on methionine production and PAS resistance was determined in M. tuberculosis. The results showed that both T120P and M172V mutations caused increased intracellular methionine concentrations and high level PAS resistance. In summary, we discovered new molecular markers and also a novel mechanism of PAS resistance in M. tuberculosis clinical isolates and broadened the understanding of the NADH-dependent MTHFR catalytic mechanism of Rv2172c in M. tuberculosis, which will facilitate the molecular diagnosis of PAS resistance and also the development of new drugs targeting Rv2172c.
期刊介绍:
Emerging Microbes & Infections is a peer-reviewed, open-access journal dedicated to publishing research at the intersection of emerging immunology and microbiology viruses.
The journal's mission is to share information on microbes and infections, particularly those gaining significance in both biological and clinical realms due to increased pathogenic frequency. Emerging Microbes & Infections is committed to bridging the scientific gap between developed and developing countries.
This journal addresses topics of critical biological and clinical importance, including but not limited to:
- Epidemic surveillance
- Clinical manifestations
- Diagnosis and management
- Cellular and molecular pathogenesis
- Innate and acquired immune responses between emerging microbes and their hosts
- Drug discovery
- Vaccine development research
Emerging Microbes & Infections invites submissions of original research articles, review articles, letters, and commentaries, fostering a platform for the dissemination of impactful research in the field.