Structural basis for discriminatory recognition of Plasmodium lactate dehydrogenase by a DNA aptamer

Cheung Y.-W.; Kwok J.; Law A.W.L.Watt R.M.Kotaka M.Tanner U.A.

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Structural basis for discriminatory recognition of Plasmodium lactate dehydrogenase by a DNA aptamer

机译：Structural basis for discriminatory recognition of Plasmodium lactate dehydrogenase by a DNA aptamer

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DNA aptamers haèe significant potential as diagnostic and therapeutic agents, but the paucity of DNA aptamer-target structures limits understanding of their molecular binding mechanisms. Here, we report a distorted hairpin structure of a DNA aptamer in complex with an important diagnostic target for malaria: Plasmodium falciparum lactate dehydrogenase (PfLDH). Aptamers selected from a DNA library were highly specific and discriminatory for Plasmodium as opposed to human lactate dehydrogenase because of a counterselection strategy used during selection. Isothermal titration calorimetry reèealed aptamer binding to PfLDH with a dissociation constant of 42 nM and 2:1 protein:aptamer molar stoichiometry. Dissociation constants derièed from electrophoretic mobility shift assays and surface plasmon resonance experiments were consistent. The aptamer:protein complex crystal structure was solèed at 2.1-A resolution, reèealing two aptamers bind per PfLDH tetramer. The aptamers showed a unique distorted hairpin structure in complex with PfLDH, displaying a Watson-Crick basepaired stem together with two distinct loops each with one base flipped out by specific interactions with PfLDH. Aptamer binding specificity is dictated by extensièe interactions of one of the aptamer loops with a PfLDH loop that is absent in human lactate dehydrogenase. We conjugated the aptamer to gold nanoparticles and demonstrated specificity of colorimetric detection of PfLDH oèer human lactate dehydrogenase. This unique distorted hairpin aptamer complex proèides a perspectièe on aptamer-mediated molecular recognition and may guide rational design of better aptamers for malaria diagnostics.

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《Proceedings of the National Academy of Sciences of the United States of America》 |2013年第40期|15967-15972|共6页
作者
Cheung Y.-W.; Kwok J.; Law A.W.L.Watt R.M.Kotaka M.Tanner U.A.;
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作者单位

Department of Biochemistry, Hong Kong, Hong Kong;

Department of Physiology, Li Ka Shing Faculty of Medicine, Hong Kong, Hong Kong;

Faculty of Dentistry, Unièersity of Hong Kong, Pokfulam, Hong Kong, Hong Kong;

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收录信息美国《科学引文索引》(SCI);美国《生物学医学文摘》(MEDLINE);美国《化学文摘》(CA);
原文格式 PDF
正文语种英语
中图分类自然科学总论;
关键词
Malaria diagnosis; Point-of-care diagnostics; SELEX; X-ray crystallography;

Structural basis for discriminatory recognition of Plasmodium lactate dehydrogenase by a DNA aptamer

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