Development of engineered E.coli for high-throughput drug screening against Malaria and Kala-azar

Student of Jawaharlal Nehru University, Delhi, Preeti Yadav won the award for a Development of engineered E.coli for high-throughput drug screening against Malaria and Kala-azar. She carried her project work under the guidance of Dr. Shailja Singh, Dr. Swati Garg, Dr. Soumya Pati.

Eukaryotic parasites increase the functional diversity of their proteome through a number of post-translational modifications (PTMs), to facilitate their survival and replication. Palmitoylation is one of the important PTM present in eukaryotic cells that modulate protein-protein interactions and plays a major role in several diseases like Cancer, Diabetes, Schizophrenia, Alzheimer’s, Malaria, Leishmaniasis, etc. In malaria parasite, Plasmodium falciparum, 10% proteome have been found to be palmitoylated that plays a crucial role in disease progression and pathogenesis, while in Leishmania donovani, we have reported that 25% of the total proteome gets palmitoylated and is involved in flagellar motility, vesicular trafficking and invasion. Plasmodium falciparum and Leishmania donovani encodes for 12 and 20 Palmitoyl acyl transferases (PATs) respectively that transfer palmitate group to target protein. Being a major contributor of parasite-diversity, palmitoylation has not been explored as a chemotherapeutic target yet due to paucity of high-throughput assays.

Prevalence of drug resistance demands immediate action to search for newer drugs and newer targets. We have developed a novel strategy involving engineered E. coli to study parasite-specific palmitoylation. E.coli is a palmitoylation machinery-null system, but our in-silico study suggested that 110 proteins of E.coli contain sites for palmitoylation. Thus, these E.coli proteins can serve as substrates for parasite-specific PATs expressed in E.coli. Parasite PATs were cloned and expressed in E.coli in the presence of PAT inhibitor and palmitoylation status of E.coli was analysed using Click chemistry. The inhibitor, 2-Bromo-palmitate that we have used here acted both as anti-malarial and anti-leishmanial as deduced by parasite growth-inhibition assay and invasion assay respectively. This is the first ex-vivo study of parasite-specific modifications in E.coli, that could be used as a robust, high-throughput screening tool for anti-protozoan drugs targeting palmitoylation, thus helping in the development of novel anti-parasitic molecules.

The Hon’ble Vice President of India, Shri M. Venkaiah Naidu awarded the Gandhian Young Technological Innovation (GYTI) Award to Preeti Yadav, Jawaharlal Nehru University, Delhi at the GYTI 2019 Awards function held at Vigyan Bhawan, New Delhi on July 06, 2019.

 

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