Free Energy Study of the Catalytic Mechanism of Trypanosoma cruzitrans-Sialidase. From the Michaelis Complex to the Covalent Intermediate

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• 作者：Gustavo Pierdominici-Sottile ; Nicole A. Horenstein ; Adrian E. Roitberg
• 刊名：Biochemistry
• 出版年：2011
• 出版时间：November 22, 2011
• 年：2011
• 卷：50
• 期：46
• 页码：10150-10158
• 全文大小：463K
• 年卷期：v.50,no.46(November 22, 2011)
• ISSN：1520-4995

文摘

Trypanosoma cruzi trans-sialidase (TcTS) is a crucial enzyme for the infection of Trypanosoma cruzi, the protozoa responsible for Chagas鈥?disease in humans. It catalyzes the transfer of sialic acids from the host鈥檚 glycoconjugates to the parasite鈥檚 glycoconjugates. Based on kinetic isotope effect (KIE) studies, a strong nucleophilic participation at the transition state could be determined, and recently, elaborate experiments used 2-deoxy-2,3-difluorosialic acid as substrate and were able to trap a long-lived covalent intermediate (CI) during the catalytic mechanism. In this paper, we compute the KIE and address the entire mechanistic pathway of the CI formation step in TcTS using computational tools. Particularly, the free energy results indicate that in the transition state there is a strong nucleophilic participation of Tyr342, and after this, the system collapsed into a stable CI. We find that there is no carbocation intermediate for this reaction. By means of the energy decomposition method, we identify the residues that have the biggest influence on catalysis. This study facilitates the understanding of the catalytic mechanism of TcTS and can serve as a guide for future inhibitor design studies.