Summary auto-generated
This study investigated how isoniazid (INH), a key tuberculosis drug, enters Mycobacterium tuberculosis cells. Researchers tracked INH uptake kinetics over minutes and examined conditions affecting transport. Initial INH uptake increased linearly with drug concentration but was unaffected by treatments blocking active transport (protonophore, ATP depletion) or by low temperature. These results indicate INH enters cells via passive diffusion rather than through protein-mediated mechanisms. However, the enzyme catalase-peroxidase (KatG) proved important for long-term accumulation of radioactivity at the plateau phase. The researchers propose that INH passively diffuses across the bacterial envelope, where KatG oxidizes it to isonicotinic acid. This oxidized form becomes trapped inside cells in its ionized state, explaining why KatG-deficient mutants show reduced drug accumulation and INH resistance. This model reconciles previous observations that KatG loss reduces labeling while clarifying that KatG facilitates drug accumulation rather than initial transport itself.
Key findings
- Isoniazid enters M. tuberculosis cells through passive diffusion, not energy-dependent active transport
- Catalase-peroxidase (KatG) is not involved in the initial transport of isoniazid but is required for intracellular accumulation of oxidized drug derivatives
- Isoniazid is oxidized by KatG to isonicotinic acid, which becomes trapped inside cells in ionized form due to the pH gradient
- KatG-deficient mutants show reduced drug accumulation because they cannot oxidize INH, explaining INH resistance in these strains
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Abstract
Initial transport kinetics of isoniazid (INH) and its uptake at the plateau were studied in Mycobacterium tuberculosis H37Rv under various experimental conditions. The initial uptake velocity increased linearly with INH concentration from 2 x 10(-6) M to 10(-2) M. It was modified neither by addition of a protonophore that abolished proline transport, nor following ATP depletion by arsenate, which inhibited glycerol uptake, two transport processes taken as controls for secondary active transport and facilitated diffusion, respectively. Microaerobiosis or low temperature (4 degrees C) were without effect on initial uptake. It is thus likely that INH transport in M. tuberculosis proceeds by a passive diffusion mechanism, and that catalase-peroxidase (KatG) is not involved in the actual transport. However, conditions inhibiting KatG activity (high INH concentration, microaerobiosis, low temperature) decrease cell radioactivity at the uptake plateau. It is proposed that INH transport occurs by passive diffusion. KatG is involved only in the intracellular accumulation of oxidized derivatives of INH, especially of isonicotinic acid, which is trapped inside cells in its ionized form. This model explains observed and previously known characteristics of the accumulation of radioactivity in the presence of [14C]INH for various species and strains of mycobacteria.