Unique properties of Plasmodium falciparum porphobilinogen deaminase
Unique properties of Plasmodium falciparum porphobilinogen deaminase
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DOI：
10.1074/jbc.m706861200
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发表时间：
2008-01-04
影响因子：
4.8
通讯作者：
Padmanaban, Govindarajan
中科院分区：
文献类型：
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作者：
Nagaraj, Viswanathan Arun;Arumugam, Rajavel;Padmanaban, Govindarajan
疟疾寄生虫中血红素生物合成的混合途径提出了位于不同区室（例如顶质体、线粒体和细胞质）的寄生虫基因组编码酶的参与。然而，关于许多这些酶的功能和定位的知识尚不可用。在这项研究中，我们证明由恶性疟原虫基因组（PfPBGD）编码的胆色素原脱氨酶具有多种独特的生化特性。使用从寄生虫膜部分纯化的 PfPBGD 以及从大肠杆菌细胞表达和纯化的缺乏 N 末端 64 个氨基酸的重组 PfPBGD (Delta PfPBGD) 进行的研究表明，这两种蛋白质都具有催化活性。令人惊讶的是，PfPBGD 催化胆色素原转化为尿卟啉原 III (UROGEN III)，表明它还具有尿卟啉原 III 合酶 (UROS) 活性，从而催化下一步。这消除了需要有一个单独的 UROS 基因的必要性，而迄今为止，该基因尚未在寄生虫基因组中注释。有趣的是，即使在热处理后，Delta PfPBGD 也会产生 UROGEN III，尽管已知其他来源的 UROS 对热敏感。基于活性位点残基的分析，创建了Delta PfPBGDL116K突变酶，该重组突变酶的比活性比Delta PfPBGD高5倍。更有趣的是，除了 UROGEN III 之外，Delta PfPBGDL116K 还催化尿卟啉原 I ( UROGEN I) 的形成，表明随着 PBGD 活性的增加，PBGD 的 UROS 活性可能会成为限速，从而导致前尿卟啉原非酶促环化为 UROGEN I。 PfPBGD 定位于顶质体，与宿主相比催化效率非常低红细胞酶。
The hybrid pathway for heme biosynthesis in the malarial parasite proposes the involvement of parasite genome-coded enzymes of the pathway localized in different compartments such as apicoplast, mitochondria, and cytosol. However, knowledge on the functionality and localization of many of these enzymes is not available. In this study, we demonstrate that porphobilinogen deaminase encoded by the Plasmodium falciparum genome (PfPBGD) has several unique biochemical properties. Studies carried out with PfPBGD partially purified from parasite membrane fraction, as well as recombinant PfPBGD lacking N-terminal 64 amino acids expressed and purified from Escherichia coli cells ( Delta PfPBGD), indicate that both the proteins are catalytically active. Surprisingly, PfPBGD catalyzes the conversion of porphobilinogen to uroporphyrinogen III (UROGEN III), indicating that it also possesses uroporphyrinogen III synthase (UROS) activity, catalyzing the next step. This obviates the necessity to have a separate gene for UROS that has not been so far annotated in the parasite genome. Interestingly, Delta PfPBGD gives rise to UROGEN III even after heat treatment, although UROS from other sources is known to be heat-sensitive. Based on the analysis of active site residues, a Delta PfPBGDL116K mutant enzyme was created and the specific activity of this recombinant mutant enzyme is 5-fold higher than Delta PfPBGD. More interestingly, Delta PfPBGDL116K catalyzes the formation of uroporphyrinogen I ( UROGEN I) in addition to UROGEN III, indicating that with increased PBGD activity the UROS activity of PBGD may perhaps become rate-limiting, thus leading to non-enzymatic cyclization of preuroporphyrinogen to UROGEN I. PfPBGD is localized to the apicoplast and is catalytically very inefficient compared with the host red cell enzyme.