Structure of Toxoplasma gondii LDH1: Active-Site Differences from Human Lactate Dehydrogenases and the Structural Basis for Efficient APAD+ Use

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• 作者：Kathryn L. Kavanagh ; Robert A. Elling ; and David K. Wilson
• 刊名：Biochemistry
• 出版年：2004
• 出版时间：February 3, 2004
• 年：2004
• 卷：43
• 期：4
• 页码：879 - 889
• 全文大小：708K
• 年卷期：v.43,no.4(February 3, 2004)
• ISSN：1520-4995

文摘

While within a human host the opportunistic pathogen Toxoplasma gondii relies heavily onglycolysis for its energy needs. Lactate dehydrogenase (LDH), the terminal enzyme in anaerobic glycolysisnecessary for NAD⁺ regeneration, therefore represents an attractive therapeutic target. The tachyzoitestage lactate dehydrogenase (LDH1) from the parasite T. gondii has been crystallized in apo form and internary complexes containing NAD⁺ or the NAD⁺-analogue 3-acetylpyridine adenine dinucleotide (APAD⁺)and sulfate or the inhibitor oxalate. Comparison of the apo and ternary models shows an active-site loopthat becomes ordered upon substrate binding. This active-site loop is five residues longer than in mostLDHs and necessarily adopts a different conformation. While loop isomerization is fully rate-limiting inprototypical LDHs, kinetic data suggest that LDH1's rate is limited by chemical steps. The importance ofcharge neutralization in ligand binding is supported by the complexes that have been crystallized as wellas fluorescence quenching experiments performed with ligands at low and high pH. A methionine thatreplaces a serine residue and displaces an ordered water molecule often seen in LDH structures providesa structural explanation for reduced substrate inhibition. Superimposition of LDH1 with human muscle-and heart-specific LDH isoforms reveals differences in residues that line the active site that increaseLDH1's hydrophobicity. These differences will aid in designing inhibitors specific for LDH1 that may beuseful in treating toxoplasmic encephalitis and other complications that arise in immune-compromisedindividuals.