Uptake and metabolism of fluorescent steroids by mycobacterial cells

Abstract

Fluorescent steroids BODIPY-cholesterol (BPCh) and 7-nitrobenzoxadiazole-4-amino-(NBD)-labeled 22-NBD-chelesterol (22NC) as well as synthesized 20-(NBD)-pregn-5-en-3β-ol (20NP) were found to undergo bioconversions by Mycobacterium tuberculosis H₃₇Rv and M. smegmatis mc² 155. The major fluorescent products were determined to be 4-en-3-one derivatives of the compounds. Degradation of NBD fluorophore was also detected in the cases of 22NC and 20NP, but neither NBD degradation nor steroidal part modification were observed for the synthesized 3-(NBD)-cholestane. Mycobacterial 3β-hydroxysteroid dehydrogenases were concluded to be responsible for the formation of the 4-en-3-one derivatives. All the compounds tested were found to cause staining both membrane lipids and cytosolic lipid droplets when incubated with mycobacteria in different manner, demonstrating ability of the steroids to reside in the compartments. The findings reveal a potential of the compounds for monitoring of steroid interactions with mycobacteria and provide information for design of new probes for this purpose.

Keywords: 22-NBD-cholesterol (PubChem CID: 127604); 3β-Hydroxysteroid dehydrogenase; Fluorescence microscopy; Fluorescent steroids; Lipid droplets; Mass-spectrometry; Menadione sodium bisulfite (PubChem CID: 23665888); Mycobacteria; Nile Red (PubChem CID: 65182).