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Mycobacterium tuberculosisis an example of a bacterial pathogen with a specialized SecA2-dependent protein export system that contributes to its virulence. Our understanding of the mechanistic basis of SecA2-dependent export and the role(s) of the SecA2 pathway in M. tuberculosispathogenesis has been hindered by our limited knowledge of the proteins exported by the pathway. Here, we set out to identify M. tuberculosisproteins that use the SecA2 pathway for their export from the bacterial cytoplasm to the cell wall. Using label-free quantitative proteomics involving spectral counting, we compared the cell wall and cytoplasmic proteomes of wild type M. tuberculosisto that of a ΔsecA2mutant. This work revealed a role for the M. tuberculosisSecA2 pathway in the cell wall localization of solute binding proteins that work with ABC transporters to import solutes. Another discovery was a profound effect of SecA2 on the cell wall localization of the Mce1 and Mce4 lipid transporters, which contribute to M. tuberculosisvirulence. In addition to the effects on solute binding proteins and Mce transporter export, our label-free quantitative analysis revealed an unexpected relationship between SecA2 and the hypoxia-induced DosR regulon, which is associated with M. tuberculosislatency. Nearly half of the transcriptionally controlled DosR regulon of cytoplasmic proteins were detected at higher levels in the ΔsecA2mutant versuswild type M. tuberculosis. By increasing the list of M. tuberculosisproteins known to be affected by the SecA2 pathway, this study expands our appreciation of the types of proteins exported by this pathway and guides our understanding of the mechanism of SecA2-dependent protein export in mycobacteria. At the same time, the newly identified SecA2-dependent proteins are helpful for understanding the significance of this pathway to M. tuberculosisvirulence and physiology.