Purification and characterization of two heme induced proteins of Mycobacterium tuberculosis

Abstract

Mycobacterium tuberculosis (Mtb), the causative agent of human tuberculosis (TB), is transmitted through aerosols and successfully replicates within alveolar macrophages by inhibiting macrophage maturation. Before the SARS-CoV-2 pandemic, Mtb had surpassed HIV/AIDS to become the leading cause of death worldwide from a single infectious agent. In 2019, 11 million people were infected with Mtb resulting in 1.3 million deaths. The WHO estimates that globally 3.3% of new cases and 18% of previously treated cases were multidrug-resistant TB (MDR-TB) in 2019. Thus, there is an imminent need to develop chemotherapeutic strategies to prevent TB disease progression. Mtb is entirely dependent on iron acquisition to colonize the human host successfully. Greater than 75% of host iron is stored in heme, making it the primary source of iron for Mtb in the human host. An overarching goal of our lab is to identify novel components of Mtb that are required for Hm iron acquisition so that we can develop novel strategies to block heme iron acquisition and starve this pathogen of an essential nutrient. Previous transcriptomic studies had identified that Mtb significantly increases expression of ppe64 and rv0125 in response to heme iron in the growth medium. The goal of this project was to clone these genes into an expression vector and attempt to purify the encoded recombinant proteins from E. coli.