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首页> 外文期刊>Biochemistry >The purine nucleoside phosphorylase from Trichomonas vaginalis is a homologue of the bacterial enzyme.
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The purine nucleoside phosphorylase from Trichomonas vaginalis is a homologue of the bacterial enzyme.

机译：来自阴道毛滴虫的嘌呤核苷磷酸化酶是细菌酶的同源物。

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摘要

Trichomonas vaginalis is a parasitic protozoan and the causative agent of trichomoniasis. Its primary purine salvage system, consisting of a purine nucleoside phosphorylase (PNP) and a purine nucleoside kinase, presents potential targets for designing selective inhibitors as antitrichomonial drugs because of lack of de novo synthesis of purine nucleotides in this organism. cDNA encoding T. vaginalis PNP was isolated by complementation of an Escherichia coli strain deficient in PNP and expressed, and the recombinant enzyme was purified to apparent homogeneity. It bears only 28% sequence identity with that of human PNP but 57% identity with the E. coli enzyme. Gel filtration showed the enzyme in a hexameric form, similar to the bacterial PNPs. Steady-state kinetic analysis of T. vaginalis PNP-catalyzed reactions gave K(m)'s of 31.5, 59.7, and 6.1 microM for inosine, guanosine, and adenosine in the nucleosidase reaction and 45.6, 35.9, and 12.3 microM for hypoxanthine, guanine, and adenine in the direction of nucleoside synthesis. This substrate specificity appears to be similar to that of bacterial PNPs. The catalytic efficiency of this enzyme with adenine as substrate is 58-fold higher than that with either hypoxanthine or guanine, representing a distinct disparity with the mammalian PNPs, which have negligible activity with either adenine or adenosine. The kinetic mechanism of T. vaginalis PNP-catalyzed reactions, determined by product inhibition and equilibrium isotope exchange, was by random binding of substrates (purine base and ribose 1-phosphate) with ordered release of the purine nucleoside first, followed by inorganic phosphate. Formycin A, an analogue of adenosine known as an inhibitor of E. coli PNP without any effect on mammalian PNPs, was shown to inhibit T. vaginalis PNP with a K(is) of 2.3 microM by competing with adenosine. T. vaginalis PNP thus belongs to the family of bacterial PNPs and constitutes a target for antitrichomonial chemotherapy.
机译：阴道毛滴虫是一种寄生虫原生动物，是滴虫的病原体。其主要的嘌呤挽救系统由嘌呤核苷磷酸化酶（PNP）和嘌呤核苷激酶组成，由于该生物体内缺乏嘌呤核苷酸的从头合成，为设计选择性抑制剂作为抗滴虫药物提供了潜在的靶标。通过互补缺乏PNP的大肠杆菌菌株分离并编码编码阴道锥虫PNP的cDNA，并表达该重组酶，使其具有明显的同质性。它与人PNP仅具有28％的序列同一性，但与大肠杆菌酶具有57％的同一性。凝胶过滤显示该酶为六聚体形式，类似于细菌PNP。稳态动力学分析阴道锥虫PNP催化的反应，核苷酶反应中肌苷，鸟苷和腺苷的K（m）为31.5、59.7和6.1 microM，次黄嘌呤的K（m）为45.6、35.9和12.3 microM，鸟嘌呤和腺嘌呤在核苷合成的方向上。该底物特异性似乎与细菌PNP相似。该酶以腺嘌呤为底物的催化效率比次黄嘌呤或鸟嘌呤的催化效率高58倍，与哺乳动物PNP明显不同，后者与腺嘌呤或腺苷的活性可忽略不计。通过产物抑制和平衡同位素交换确定的阴道锥虫PNP催化反应的动力学机制是通过底物（嘌呤碱和1-磷酸核糖）的无规结合，首先有序释放嘌呤核苷，然后是无机磷酸盐。 Formycin A是腺苷的类似物，被称为大肠杆菌PNP的抑制剂，对哺乳动物PNP没有任何影响，已显示可通过与腺苷竞争而抑制K.is为2.3 microM的阴道T. PNP。因此，阴道锥虫PNP属于细菌PNP家族，并构成抗滴虫化学疗法的靶标。

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