Mycobacterium tuberculosis (Mtb) remains the most deadly bacterial pathogen, and rampant drug resistance requires renewed efforts to find new and better therapies. At the core of Mtb’s success as a pathogen lies its ability to sense and adapt to host cues, in particular through sensor protein kinase signaling. The Grundner lab maps these signaling pathways and determines their role in Mtb physiology. These studies provide fundamental insight into Mtb biology and identify new targets for therapeutic interference. Because the Mtb Ser/Thr kinases are related to eukaryotic kinases, we repurpose the extensive pharmaceutical resources developed for targeting human kinases for use against Mtb. A major bottleneck in Mtb research on every level is the large number of genes with unknown function in the Mtb genome. Determining protein function is currently exceedingly slow and proceeds one protein at a time. To scale functional annotation to the level of other -omics approaches, we use activity-based protein profiling combined with mass spectrometry towards high-throughput identification of functions for these unknown proteins. These new tools allow probing of even the most divergent enzyme space and provide a more complete understanding of the genomic dark matter of Mtb.