Tarique Mohammed, Satsangi Akash Tripathi, Ahmad Moaz, Singh Shailja, Tuteja Renu
Malaria Group, International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Marg, New Delhi 110067, India.
Mol Biochem Parasitol. 2012 Feb;181(2):153-61. doi: 10.1016/j.molbiopara.2011.10.012. Epub 2011 Nov 4.
Malaria is one of the most important infectious diseases in many regions around the world including India. Plasmodium falciparum is the cause of most lethal form of malaria while Plasmodium vivax is the major cause outside Africa. Regardless of considerable efforts over the last many years there is still no commercial vaccine against malaria and the disease is mainly treated using a range of established drugs. With time, the malaria parasite is developing drug resistance to most of the commonly used drugs. This drug resistance might be due to defective mismatch repair in the parasite. Previously we have reported that the P. falciparum genome contains homologues to most of the components of mismatch repair (MMR) complex. In the present study we report the detailed biochemical characterization of one of the main component of MMR complex, MLH, from P. falciparum. Our results show that MLH is an ATPase and it can incise covalently closed circular DNA in the presence of Mn(2+) or Mg(2+) ions. Using the truncated derivatives we show that full length protein MLH is required for all the enzymatic activities. Using immunodepletion assays we further show that the ATPase and endomuclease activities are attributable to PfMLH protein. Using immunofluorescence assay we report that the peak expression of MLH in both 3D7 and Dd2 strains of P. falciparum is mainly in the schizont stages of the intraerythrocytic development, where DNA replication is active. MMR also contributes to the overall fidelity of DNA replication and the peak expression of MLH in the schizont stages suggests that MLH is most likely involved in correcting the mismatches occurring during replication. This study should make a significant contribution in our better understanding of DNA metabolic processes in the parasite.
疟疾是包括印度在内的世界许多地区最重要的传染病之一。恶性疟原虫是最致命形式疟疾的病因，而间日疟原虫是非洲以外地区疟疾的主要病因。尽管在过去许多年里付出了巨大努力，但仍然没有针对疟疾的商业疫苗，该疾病主要使用一系列既定药物进行治疗。随着时间的推移，疟原虫对大多数常用药物产生了耐药性。这种耐药性可能是由于寄生虫中错配修复缺陷所致。此前我们报道过，恶性疟原虫基因组包含错配修复（MMR）复合体大多数组分的同源物。在本研究中，我们报道了来自恶性疟原虫的MMR复合体主要组分之一MLH的详细生化特性。我们的结果表明，MLH是一种ATP酶，在存在Mn(2+)或Mg(2+)离子的情况下，它可以切割共价闭合环状DNA。使用截短衍生物，我们表明全长蛋白MLH对所有酶活性都是必需的。通过免疫去除分析，我们进一步表明ATP酶和核酸内切酶活性归因于PfMLH蛋白。通过免疫荧光分析，我们报道在恶性疟原虫的3D7和Dd2菌株中，MLH的峰值表达主要在红细胞内发育的裂殖体阶段，此时DNA复制活跃。MMR也有助于DNA复制的整体保真度，并且MLH在裂殖体阶段的峰值表达表明MLH很可能参与纠正复制过程中发生的错配。这项研究应该会对我们更好地理解寄生虫中的DNA代谢过程做出重大贡献。
