Mycobacterium tuberculosis is an air borne, facultative intracellular bacterial pathogen that resides in the phagosome of host cells. Virulence of M. tuberculosis is related to its abilities to respond to environmental cues encountered during infection and reprogram its metabolism to adapt to them. Nutrients derived from the host are key factors contributing to shape the carbon metabolism of M. tuberculosis during infection. This metabolic shift involves activation of fatty acid and cell wall lipids metabolism of the bacterium; however, the mechanistic interplay between them is undefined. In this study, to understand the metabolic adaptation on propionyl-CoA 3carbon product of cholesterol in M. tuberculosis, the propionate detoxification and methyl-branched (MB) lipid synthesis pathways were investigated. The data presented here shows that excess propionyl-CoA was toxic to M. tuberculosis, as the propionylCoA inhibited pyruvate dehydrogenase (PDH) and acetate or fatty acid rescued the toxicity through providing acetyl-CoA the product of PDH. A mechanistic insight was revealed by metabolic labeling with radioactive propionate and fatty acids: the given fatty acids facilitate propionyl-CoA incorporation into a key MB lipid PDIM by serving as acyl-primers required for its biosynthesis. Additionally, to further define genes required for propionate and fatty acid metabolism an approach exploiting propionate toxicity and TraSH analysis was employed. The data from the genetic profiling by TraSH confirms our model for the fatty acids rescue from the propionate toxicity and suggests putative roles for uncharacterized genes in MB lipids synthesis and fatty acid metabolism. This model was also validated by using lipid droplet loaded macrophage where M. tuberculosis exploited fatty acids from the host lipid droplet to synthesis PDIM resulting in reduction of the propionate stress. Data presented in this work demonstrates that the propionyl-CoA processing is a significant problem for survival in the host cell, and that the routing of propionyl-CoA into MB cell wall lipids plays an important role for limiting the metabolic stress derived from propionate. And, the data also demonstrates that the fatty acids from lipid droplets in the host macrophage may provide a well-balanced diet that retains appropriate level of acetyl-CoA and propionyl-CoA.