Fig 4. The transmembrane domain of Rv1625c is essential for complete degradation of cholesterol and the catalytic domain of Rv1625c is required for V-59 activity.

(A) Catabolic release of ¹⁴CO₂ from [4-¹⁴C]-cholesterol in WT, ΔRv1625c, Comp_Full, and Comp_D204 strains treated with V-59 (10 μM) or DMSO vehicle control. Data are from two experiments with three technical replicates, normalized to OD and quantified relative to WT treated with DMSO. Shown as means ± SD (**P < 0.01, ***P < 0.001, Two-way ANOVA with Tukey’s multiple comparisons test). (B) TLC comparing [4-¹⁴C]-cholesterol-derived metabolites extracted from supernatants of Mtb. Image is representative of two experiments. Equivalent counts were spotted per lane. “Chol.” = [4-¹⁴C]-cholesterol.