Figure 2.

The cascade of PhenoTarget screening for identifying lead compounds and target proteins is shown. The Nature Bank (NB) lead-like enhanced (LLE) fraction library was established following the procedure described by Camp et al. [14]. A high throughput phenotypic screening of 202,983 NB LLE fractions against M. tuberculosis H37Rv was initially performed. Active fractions with an MIC value of less than 6.1 µge/µL were identified and chosen for protein screening against a panel of 37 putative anti-TB targets from Mycobacteria species. To lower sample consumption, especially protein, nine active fractions were pooled (Pool Fractions 1 to 40) and incubated with each of the target proteins. Native mass spectrometry was then used to identify free target protein (P, blue) and protein-ligand (P-L) complexes. The mass shift between the P (black) and the P-L (red) peaks provided the molecular weight of the bound ligand (L, purple) and this facilitated the isolation and identification of the active compound.