The Catalytic Mechanism of Pdx2 Glutaminase Driven by a Cys-His-Glu Triad: A Computational Study

• Type: Article in International Scientific Journal
• Date: 2021

Authors

Pina, AF (Author) Other; Sergio Filipe Sousa (Author) FCUP; Cerqueira, NMFSA (Author) Other

Journal

Title: ChemBioChem

ISSN: 1439-4227

Publisher: Wiley-Blackwell

Indexing

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Other information

Authenticus ID: P-00V-TEA

DOI: 10.1002/cbic.202100555

Abstract (EN): The catalytic mechanism of Pdx2 was studied with atomic detail employing the computational ONIOM hybrid QM/MM methodology. Pdx2 employs a Cys-His-Glu catalytic triad to deaminate glutamine to glutamate and ammonia - the source of the nitrogen of pyridoxal 5'-phosphate (PLP). This enzyme is, therefore, a rate-limiting step in the PLP biosynthetic pathway of Malaria and Tuberculosis pathogens that rely on this mechanism to obtain PLP. For this reason, Pdx2 is considered a novel and promising drug target to treat these diseases. The results obtained show that the catalytic mechanism of Pdx2 occurs in six steps that can be divided into four stages: (i) activation of Cys(87), (ii) deamination of glutamine with the formation of the glutamyl-thioester intermediate, (iii) hydrolysis of the formed intermediate, and (iv) enzymatic turnover. The kinetic data available in the literature (19.1-19.5 kcal mol(-1)) agree very well with the calculated free energy barrier of the hydrolytic step (18.2 kcal.mol(-11)), which is the rate-limiting step of the catalytic process when substrate is readily available in the active site. This catalytic mechanism differs from other known amidases in three main points: i) it requires the activation of the nucleophile Cys(87) to a thiolate; ii) the hydrolysis occurs in a single step and therefore does not require the formation of a second tetrahedral reaction intermediate, as it is proposed, and iii) Glu(198) does not have a direct role in the catalytic process. Together, these results can be used for the synthesis of new transition state analogue inhibitors capable of inhibiting Pdx2 and impair diseases like Malaria and Tuberculosis.

Language: English

Type (Professor's evaluation): Scientific

No. of pages: 13