Uptake and Binding of ¹⁴ C-Ethambutol by Tubercle Bacilli and the Relation of Binding to Growth Inhibition

Abstract

Studies were designed to characterize ethambutol uptake by Mycobacterium tuberculosis (H37Ra) and to relate uptake to the time-dependent, concentrationindependent nature of growth inhibition by ethambutol. When cells grown aerated at 37 C in Sauton medium were exposed for 7 hr to 0.2, 0.5, 1.0, 2.5, and 5.0 ,ug of 14C-ethambutol per ml, uptake increased with time and was a linear function of concentration. The process was inhibited at 22 C. Studies with chloramphenicol, sodium azide, and 2,4-dinitrophenol indicated that uptake is independent of requirements for protein synthesis and energy. The organism did not accumulate ethambutol against a concentration gradient. It can be concluded that ethambutol enters the cells in a passive manner. Kinetic studies of 14C loss from tubercle bacilli pretreated with labeled drug suggested the existence of two ethambutol fractions within the cell: a highly variable labile pool and a second fraction that is small and quite firmly bound. Levels of cell-bound drug may be independent of total uptake, but this possibility was not established unequivocally. Definitive evidence showing identity in the concentrations of bound drug regardless of total uptake could explain the apparent discrepancy between concentration-dependent uptake and concentration-independent growth inhibition.

Tryptophan Uptake By Mycobacterium tuberculosis H 37 Rv: Effect of Rifampin and Ethambutol

“…Ethambutol may be blocking the uptake like the latter compounds. Earlier studies with M. tuberculosis H37Ra (1) (Fig 9).…”

Proteomic Analysis of the Mycobacterium tuberculosis Outer Membrane for Potential Implications in Uptake of Small Molecules

“…The aqueous phases from both rounds of extraction were pooled and subjected to drying by a speed vacuum concentration. A total of 600 μL of NMR buffer was added to the lyophilized M. tuberculosis 13 C, 15 N, 31 P, 2 H) installed on an AVANCE III HD Ascend Bruker NMR spectrometer with a magnetic field of 14.1 T was used to record all the NMR data at 298 K. Noesygppr1d pulse sequence was selected from Bruker library; it is a water suppression pulse sequence and uses water presaturation in the form of RD-G 1 -90°-t-90°-t m -G 2 -90°-ACQ, where RD is the relaxation delay between two successive scans of 5 s, t is the short time delay of ∼3 μs, 90°means a 90°radio frequency pulse, t m means the mixing time (100 ms), and ACQ means the acquisition time (6.95 s). Each spectrum was recorded for 64 scans with 16 dummy scans into a spectral width of 7200 Hz and 32,000 data points.…”

Free Energy Barriers for Passive Drug Transport through the Mycobacterium tuberculosis Outer Membrane: A Molecular Dynamics Study

“…Among these compounds, INH, PZA, EMB, and RIF are the classical first-line antibiotics that have been used in clinical practice for a long time, while BDQ, PRET, and MOX are relatively new and promising drugs. For the INH, PZA, EMB, RIF, and BDQ molecules, there is experimental evidence or reasonable assumptions of the passive diffusion mechanism of their transport into Mtb cells [15,[26][27][28][29][30][31]. The exception is PZA, which can also cross the cell wall by several other mechanisms.…”