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6th Edition of World Congress on Infectious Diseases

June 24-26, 2024 | Paris, France

June 24 -26, 2024 | Paris, France
Infection 2024

Faryal Ashraf

Speaker at World Congress on Infectious Diseases 2024 - Faryal Ashraf
International Center for Chemical and Biological Sciences, Pakistan
Title : Progress in the field of structural biology in Pakistan: Calculation of 1st solution NMR structure


A robust progress in the field of structural biology has allowed the scientists to see the architectural beauty of the macromolecules along with their dynamics. The variety of resolution comprehends by different techniques allow to observe the changes in the macromolecules over time as well as how they interact with other partners. These advancements resulted in deeper understanding of biomolecules and their functions. Therefore, playing significant role in health, diseases and mechanism manifested drug discovery.

This study presents a first solution NMR structure calculated in Pakistan. The drug target selected for this purpose was purS protein. The purS protein is a subunit of phosphoribosylformyglycinamidine synthase. This enzyme catalyzes the fourth step reaction of denovo purine biosynthesis (Fig-1). Hence, it is a key player of DNA or RNA synthesis.

The purS subunit was cloned, expressed and purified using in-house facility. The vancomycin resistant Staphylococcus aureus (ATCC 700699) was used as model organism for gene target. However, the E. coli DH5α strain was used to clone the gene and E. coli BL21(DE3) strain was used for expression of protein. The protein purification was carried out using affinity column on Fast Protein Liquid Chromatography (FPLC). The 6X His-tag used for purification was then removed using TEV protease. The 2D and 3D NMR experiments were acquired on Bruker Avance III HD 800 MHz spectrometer equipped with TCI-HCN cryogenically cooled probe. The CARA and SPARKY software was used for the backbone assignments. Whereas, final structure calculations were done using the CYANA 3.98.15 software package. The interaction of purS subunit with Glutamine and ATP molecule was also assessed by Chemical Shift Perturbation.

The key findings include the refined solution NMR structure of purS subunit (PDB ID: 8J3N) and the subunit does not bind with glutamine whereas, show affinity with the ATP molecule. The solution NMR structure of PurS subunit showed that the N-, and C-terminal residues are located in the core of the protein and stabilized by hydrogen bond and electrostatic interactions. While the core of protein is surrounded by α-helix, and 310-helix. The structure of purS subunit possess novel fold as no DALI hit matches with the submitted query.


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