ABSTRACT
While the new coronavirus has turned our lives upside down causing millions of deaths, the historically known tuberculosis (TB) disease caused by Mycobacterium tuberculosis (MTB) was responsible for the loss of approximately 1.5 million lives alone in 2021. New anti- TB drugs are in an urgent need. A promising target is dUTPase, an enzyme preventing uracil incorporation into DNA. It is present in all multicellular species and in most microbes. Abolition of its activity potentially leads to DNA double strand breaks and cell death. Therefore, species-specific inhibition of MTB dUTPase may be a successful way of TB disease treatment. Currently no species-specific dUTPase inhibitor exists, but an interaction partner, protein Stl shows significantly different ability to inhibit dUTPase homologues from various species. We use Stl as a model to understand how species- specific differences in dUTPase structure may be harnessed in future inhibitor development. A remarkable species-specific characteristic of MTB dUTPase is a small surface sequence loop playing no direct role in enzyme activity but being essential for mycobacterial survival in a yet unknown way. What is the exact structural background of MTB dUTPase-Stl interaction? For this reason, we have crystallized a complex of MTB dUTPase and a truncated Stl protein mutant. And how the loop sequence may affect the MTB dUTPase protein structure on its own? For this answer, we obtained another X-ray diffraction dataset of a loop-lacking mutant of MTB dUTPase with 1.3 Å resolution. Surprisingly, electron density of the flexible C-terminal “arm” segment of the mutant dUTPase was missing from our dataset, contrary to the already crystallized wildtypeMTB dUTPase structures. We postulate that the loop sequence may restrict conformational flexibility of the dUTPase “arm”, making it more inhibitable by Stl compared to the loop-lacking mutant, as we know from our comparative steady-state enzyme activity inhibition measurements.