Structural and dynamics studies of FKBP12 from Different Microorganisms: Biological Target for Inhibitors Against Tuberculosis and Neglected Diseases

Neglected Tropical Diseases (NTDs) are found in tropical and subtropical world regions and constitute a substantial public health problem. The tuberculosis epidemic annually affects about 10 million people worldwide. Bacterial resistance and the lack of more effective therapy have become a considerable concern. Thus, the investigation of new molecular targets and the development of more potent drugs against these diseases is crucial. FKBP12 proteins, an essential peptidyl-prolil-cis-trans-isomerase to numerous disease-causing microorganisms, are reported as potential biological targets. They differ about 40% in their primary sequence compared to their human ortholog. This work aims to study the structure and dynamics of FKBP12 from different microorganisms and compare it to Homo sapiens ortholog. Prospection studies using the primary sequence of human FKBP12 were done using bioinformatics tools. FKBP12 from Mycobacterium tuberculosis (MtFKBP12) and Leishmania infantum (LiFKBP12) were selected. Previously, the group performed structural, dynamics, and interaction studies with MtFKBP12. LiFKBP12 were cloned into pET28a and transformed in E. coli strains. Expression tests were performed using different parameters and were monitored by 15% SDS-PAGE. Results and discussion: LiFKBP12 best expression condition was achieved using BL21(DE3) strain and induction with 0.5 mM of IPTG (OD600>1.0), at 18oC for 4 hours. Purification methods were applied using two steps: affinity and size-exclusion chromatography. His-tag was cleaved with TEV protein (1:10) using dialysis for 16h at 4oC. LiFKBP12 protein will be isotopically labeled with 15N and 13C to perform all experiments required for NMR assignment and dynamics studies. These studies will bring essential information about specific regions of FKBPs that will be important to the studies of fragment screening for the search for new compounds against Tuberculosis and NTDs.