Rutgers New Jersey Medical School
Microbiology, Biochemistry & Molecular Genetics
Exploring membrane proteins involved in polymyxin resistance using cryo-EM
University of Kentucky
April 2018
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December 2020
Pharmaceutical Sciences
Passionate about the chemistry of macro-molecular interactions and disease biology. Currently, I am exploring new strategies to combat multiple drug resistance in tuberculosis as a Post-Doctoral Scholar in University of Kentucky, Kentucky, USA (2018-2020). I use X-ray crystallography to understand enzyme-inhibitor interactions at atomic level coupled with biochemical assays. A structure based drug design approach is used to develop better inhibitors.
The work involves cloning, expression, purification, biochemical assays, crystallization and structure determination.
Indian Institute of Technology, Guwahati
Biosciences and Bioengineering
The work involved various techniques - cloning, expression, purification, characterization using various biochemical and biophysical assays, site-directed mutagenesis, crystallization, RNA extraction, single nucleotide ladder generation to characterize the RNase activity, fluorescent labeling, tryptophan fluorescence quenching assays.
Project: Investigations on the maturation of CRISPR RNA in type I-C CRISPR-Cas system.
Result: Investigations led to the identification of the novel modes opted by the Cas proteins to recognize and process the substrate, involved in type I-C CRISPR variant’s adaptive immunity pathway. The distinctive feature found is the presence two endonucleases viz., Cas5d and Csd1 showing metal dependent plasticity in the activity. Intriguingly, a metal dependent activity switching was identified, exposing the beautiful versatility of the system – where a metal independent RNase takes over a promiscuous metal dependent DNase role in adaptive immunity pathway. Remarkably, the CRISPR repeat RNA processing proceeds with specificity forming specific products, while the DNase activity is apparently non-specific and the degradation occurs till the single nucleotide level. Among the two nucleases identified, Csd1 was found to be biased towards double stranded DNA, while Cas5d showed no such preference and cleaved all forms of DNA in presence of metal. Further, Cas5d and Csd1 assembled together with inactive protein, Csd2 to form the Cascade (CRISPR associated complex for antiviral defense) of type I-C. In in vivo, Cascade possesses pre-crRNA processing ability but no apparent DNase activity.
Article is published in Nucleic Acid Research.
